Photodynamic reactions using high-intensity red LED promotes gingival wound healing by ROS induction.

Photodynamic therapy is a treatment that combines a light source with a photosensitizer. LEDs have attracted considerable attention in clinical dentistry because they are inexpensive and safe to use. Although the interaction between photosensitizers and LEDs in dental practice is effective for treating periodontal disease by killing periodontopathic bacteria, little is known about the effects of LEDs on human gingival fibroblasts (HGnFs), which play an important role in gingival wound healing. In this study, we investigated the effects of high-intensity red LED irradiation on HGnFs after the addition of methylene blue (MB), one of the least harmful photosensitizers, on wound healing and reactive oxygen species (ROS) production induced by photodynamic reactions. We found that irradiation of MB with high-intensity red LED at controlled energy levels promoted cell proliferation, migration, and production of wound healing factors. Furthermore, ROS production by a photodynamic reaction enabled the translocation of phosphorylated Grb2-associated binder-1, activating Extracellular signal-regulated kinase 1/2 and c-Jun N-terminal kinase signals. Our findings suggest that proper control of ROS production has a beneficial effect on gingival fibroblasts, which constitute periodontal tissue, from the perspective of gingival wound healing.

Essential amino acid starvation induces cell cycle arrest, autophagy, and inhibits osteogenic differentiation in murine osteoblast.

This study investigates the effects of essential amino acid (EAA) starvation on murine osteoblasts cells and the underlying mechanisms. We performed and observed the cell proliferation, autophagy, and osteogenic differentiation under deprivation of EAA in vitro. The results showed that EAA starvation resulted in cell cycle arrest via phosphorylation of the MAPK signaling pathway, leading to inhibition of cell proliferation and osteogenic differentiation. Additionally, the LKB1-AMPK signaling pathway was also found to be phosphorylated, inducing autophagy. These findings highlight the significant role of EAA in regulating cellular processes. Furthermore, this study contributes to our understanding of the effects of nutrient deprivation on cellular physiology and may aid in the development of novel therapeutic strategies for diseases associated with amino acid metabolism.

High-Intensity Red Light-Emitting Diode Irradiation Suppresses the Inflammatory Response of Human Periodontal Ligament Stem Cells by Promoting Intracellular ATP Synthesis.

Periodontitis is an inflammatory lesion in the periodontal tissue. The behavior of human periodontal ligament stem cells (hPDLSCs), which play an important role in periodontal tissue regeneration, is restricted by the influence of inflammatory mediators. Photobiomodulation therapy exerts anti-inflammatory effects. The purpose of this study was to investigate the effects of light-emitting diode (LED) irradiation on the inflammatory responses of hPDLSCs. The light source was a red LED (peak wavelength: 650 nm), and the total absolute irradiance was 400 mW/cm2. The inflammatory response in hPDLSCs is induced by tumor necrosis factor (TNF)-α. Adenosine triphosphate (ATP) levels and pro-inflammatory cytokine (interleukin [IL]-6 and IL-8) production were measured 24 h after LED irradiation, and the effects of potassium cyanide (KCN) were investigated. LED irradiation at 6 J/cm2 significantly increased the ATP levels and reduced TNF-α-induced IL-6 and IL-8 production. Furthermore, the inhibitory effect of LED irradiation on the production of pro-inflammatory cytokines was inhibited by KCN treatment. The results of this study showed that high-intensity red LED irradiation suppressed the TNF-α-stimulated pro-inflammatory cytokine production in hPDLSCs by promoting ATP synthesis. These results suggest that high-intensity red LED is a useful tool for periodontal tissue regeneration in chronically inflamed tissues.

Irradiation by high-intensity red light-emitting diode enhances human bone marrow mesenchymal stem cells osteogenic differentiation and mineralization through Wnt/ß-catenin signaling pathway.

Photobiomodulation therapy (PBMT) using a light-emitting diode (LED) has been employed for various photomedicine studies. The aim of this study was to determine the effects of a high-intensity red LED on the proliferation and osteogenic differentiation of human bone marrow mesenchymal stem cells (BMSCs) and the related mechanism. BMSCs were subjected to high-intensity red LED (LZ1-00R205 Deep Red LED) irradiations for 0 to 40 s with energy densities ranging from 0 to 8 J/cm2. The distance from the LED to the cell layer was 40 mm. The spot size on the target was 4 cm2. Cell proliferation was measured at 3, 24, 48, and 72 h. The effects of LED irradiation on osteogenic differentiation and mineralization were examined with a particular focus on the Wnt/ß-catenin signaling pathway. The high-intensity red LED irradiations did not alter BMSC proliferation after 72 h. LED exposure of 6 J/cm2 (30 s) led to significant enhancements of osteogenic differentiation and mineralization. Additionally, the high-intensity LED irradiation induced activation of Wnt/ß-catenin. The effects of the high-intensity LED irradiation on BMSC osteogenic differentiation and mineralization were suppressed by treatment with the Wnt/ß-catenin inhibitor XAV939. P < 0.05 was considered significant. The results indicate that high-intensity red LED irradiation increases BMSC osteogenic differentiation and mineralization via Wnt/ß-catenin activation. Therefore, short duration irradiation with a portable high-intensity LED may be used as a potential approach in hard tissue regeneration therapy.

High-power, red-light-emitting diode irradiation enhances proliferation, osteogenic differentiation, and mineralization of human periodontal ligament stem cells via ERK signaling pathway.

BACKGROUND: Light-emitting diode (LED) is attracting attention as a new light source for phototherapy. However, its effects on periodontal tissue regeneration remain unknown. The aim of this study was to examine the effects of high-power, red LED irradiation on human periodontal ligament stem cells (PDLSCs), which play an important role in periodontal tissue regeneration. METHODS: PDLSCs were derived from adult human third molars. The light source was red LED (peak wavelength: 650 nm). Energy densities ranging from 0 to 10 J/cm2 were tested to determine the optimal dose. PDLSC proliferation was measured using two parameters: live cell protease and ATP levels. After the cells were induced to differentiate, the effect of LED irradiation on osteogenic differentiation and mineralization was examined, with particular focus on the extracellular signal-regulated kinase (ERK)1/2 signaling pathway using an ERK inhibitor (PD98059). RESULTS: LED irradiation at 8 J/cm2 led to a significant increase in PDLSC proliferation and enhanced Runx2 and Osterix mRNA expression, Alkaline phosphatase activity, procollagen type I C-peptide and osteocalcin production, calcium deposition, and alizarin red S staining. In addition, LED induced the activation of ERK1/2, and the effects of LED on PDLSC proliferation, differentiation, and mineralization could be suppressed by treatment with PD98059. CONCLUSIONS: The results of this study show that 650-nm high-power, red, LED irradiation increases PDLSCs proliferation, and osteogenic differentiation and mineralization, mediated by ERK1/2 activation. These findings suggest that LED may be a useful tool for periodontal tissue regeneration.

High Glucose Concentrations Suppress the Proliferation of Human Periodontal Ligament Stem Cells and Their Differentiation Into Osteoblasts.

BACKGROUND: Diabetes mellitus (DM) is a major risk factor for periodontal disease and affects various cellular functions. Periodontal ligament stem cells (PDLSCs) play an important role in periodontal tissue regeneration; however, the effect of hyperglycemia on PDLSCs is unclear. The aim of this study is to investigate whether hyperglycemia affects periodontal tissue regeneration, using human PDLSCs and high-glucose medium as a model of DM. METHODS: PDLSCs were obtained from healthy adult human mandibular third molars. Cell proliferation, osteoblastic differentiation, and proinflammatory cytokine expression were investigated by culturing PDLSCs in media supplemented with four different glucose concentrations representative of control patients (5.5 mM), patients with postprandial or controlled DM (8.0 mM), and patients with uncontrolled DM (12.0 and 24.0 mM). The molecular effects of hyperglycemia on PDLSC physiology were examined with a focus on the nuclear factor (NF)-(κB signaling pathway. The involvement of NF-κB was investigated with a specific NF-κB inhibitor in PDLSCs under hyperglycemic conditions. RESULTS: High glucose levels inhibited PDLSC proliferation and differentiation into osteoblasts but induced NF-κB activation and subsequent interleukin (IL)-6 and IL-8 expression. Treatment with an NF-κB inhibitor rescued the defects in cell proliferation and osteoblastic differentiation and inhibited the IL-6 expression caused by the high-glucose environment. CONCLUSION: The results of this study demonstrate that hyperglycemia inhibits human PDLSC proliferation and osteoblastic differentiation.

Small bowel perforation from a thermal burn caused by contact with the end of a laparoscope during ovarian cystectomy.

Although laparoscopic surgery now replaces many gynecologic laparotomy procedures, serious complications unique to laparoscopy may occur, including vascular or bowel injury. In most cases of bowel injury during laparoscopy, the laparoscopic instruments that cause injury are the trocar, Veress needle, grasping forceps or scissors, electrocoagulator, or laser. We report a rare case of small bowel perforation after a thermal burn caused by contact with the end of the scope during laparoscopic ovarian cystectomy. Burns and perforations of the small bowel during laparoscopy are rare complications preventable by familiarity with the physical properties of the laparoscopic instruments.

Tumor necrosis factor-alpha induced the release of interleukin-6 from endometriotic stromal cells by the nuclear factor-kappaB and mitogen-activated protein kinase pathways.

OBJECTIVE: To examine the involvement of nuclear factor-kappaB (NF-kappaB) and mitogen-activated protein kinase (MAPK) in the induction of interleukin-6 (IL-6) by tumor necrosis factor-alpha (TNF-alpha) in endometriotic stromal cells (ESC). DESIGN: Prospective study. SETTING: Department of Obstetrics and Gynecology, Tottori University Hospital, Yonago, Japan. PATIENT(S): Twelve patients who underwent laparoscopic surgery. INTERVENTION(S): Endometriotic stromal cells were obtained from chocolate cyst linings of the ovary. MAIN OUTCOME MEASURE(S): We determined the effect of TNF-alpha on the production of IL-6 and the effect of inhibitors for NF-kappaB and the MAPK pathway on IL-6 production using ELISA. Western blottings and electrophoretic mobility shift assays were used to detect activation of NF-kappaB and extracellular signal-regulated kinase 1/2 (ERK1/2). RESULT(S): The addition of TNF-alpha (0.1 ng/mL) significantly increased IL-6 protein in endometriotic stromal cells. Western blottings and electrophoretic mobility shift assays revealed that incubation with TNF-alpha induced degradation of inhibitor kappaB (I kappaB) and expression of phosphorylated ERK1/2. The NF-kappaB inhibitor (TPCK) and MAPK inhibitor (U0126) blocked the TNF-alpha-induced IL-6 expression. Electrophoretic mobility shift assay revealed that U0126 attenuated activator protein-1 (AP-1) activation induced by TNF-alpha. CONCLUSION(S): These findings demonstrate that NF-kappaB and AP-1 activation is critical for TNF-alpha-induced IL-6 expression in endometriotic stromal cells. Novel therapeutic modalities targeting these molecules may be possible in the near future.

A combination of interleukin-6 and its soluble receptor impairs sperm motility: implications in infertility associated with endometriosis.

BACKGROUND: We previously reported that the level of interleukin (IL)-6 is increased in the peritoneal fluid of women with endometriosis. This study was undertaken to assess the effects of IL-6 and soluble IL-6 receptor (sIL-6R) on in vitro sperm motility. METHODS: Sperm (n = 20) were cultured with IL-6 or sIL-6R, or with a combination of both. After 24 h cultures, sperm motility was evaluated using a computer-assisted semen analysis system. Gene and protein expressions of IL-6, IL-6 receptor (IL-6R), and glycoprotein 130 (gp130) were examined in sperm by RT-PCR analysis and western blot analysis. RESULTS: Addition of IL-6 or sIL-6R individually to the culture media had no affect on sperm motion. However, adding a combination of IL-6 and sIL-6R dose-dependently reduced the percentage of motile and rapidly moving sperm. Adding anti-IL-6R antibody abolished these adverse effects. Sperm expressed the gp130 gene and protein, but not IL-6 or IL-6R. CONCLUSIONS: A combination of IL-6 and sIL-6R may be associated with gp130 expressed in the sperm and reduce sperm motility. IL-6 and sIL-6R may contribute to the pathogenesis of endometriosis-associated infertility.

Activation of mitogen-activated protein kinase pathway by keratinocyte growth factor or fibroblast growth factor-10 promotes cell proliferation in human endometrial carcinoma cells.

Fibroblast growth factors (FGFs) exert diverse effects resulting from their interaction with cognate receptors on target cells. Our current study was designed to examine the local production and action of two specific stromal-epithelial cell mediatory factors, keratinocyte growth factor (KGF) and FGF-10, in human endometrial carcinoma cells. The RT-PCR method was used to determine gene expression of KGF, FGF-10, and KGF receptor in human endometrial carcinoma cells (HEC-1) and human endometrial stromal cells. KGF mRNAs were expressed in both of these cell types. On the other hand, FGF-10 mRNA was detected only in the endometrial stromal cells, and KGF receptor mRNA was observed in the HEC-1 cells. The novel finding of the present study is that KGF is expressed in carcinoma cells and FGF-10 is expressed in human endometrial stromal cells. The distinct phosphorylation of ERK-1 and -2 (ERK1/2), which are members of the MAPK family, was observed when HEC-1 cells were treated with KGF or FGF-10. KGF and FGF-10 could induce the prompt phosphorylation of ERK1/2 and consequently stimulate DNA synthesis. KGF and FGF-10 did not activate the phosphorylation of Akt, protein kinase C, or signal transducer and activator of transcription-3. Blocking the MAPK pathway with the specific methyl ethyl ketone 1/2 inhibitor (U0126) completely neutralized the enhancement of cell proliferation induced by KGF and FGF-10. In addition, KGF and FGF-10 activated expressions of downstream nuclear transcription factors, such as Elk-1 and c-myc, but not c-fos. These results demonstrate for the first time that KGF and FGF-10 are capable of stimulating the growth of endometrial carcinoma cells via activating MAPK pathway through autocrine/paracrine fashion.

Interleukin-8 gene and protein expression are up-regulated by interleukin-1beta in normal human ovarian cells and a granulosa tumor cell line.

OBJECTIVE: To evaluate the expression, regulation, and role of interleukin (IL)-8 in human ovary. DESIGN: Prospective study. SETTING: University hospital. PATIENT(S): Sixteen premenopausal women. INTERVENTION: Follicular fluid and granulosa lutein cells (GLCs) were collected during IVF cycles. Ovarian stromal and theca cells were obtained from women underwent surgery. KGN cells, the human granulosa cell tumor cell line, were also used. MAIN OUTCOME MEASURES: The levels of IL-8 and IL-1beta in follicular fluid and IL-8 protein production were determined using ELISA. Interleukin-8 and IL-8 receptor gene expression in ovarian cells and the effect of IL-8 on the proliferation of stromal cells were determined. The expression of pIkappaB was evaluated by Western blot, and the effect of NF-kappaB inhibitor APDC was examined by Northern blot analysis and ELISA in KGN cells. The levels of IL-8 and IL-1beta in follicular fluid; each concentration and the volume showed a positive correlation. Reverse transcription polymerase chain reaction showed the presence of IL-8 mRNA in all ovarian cells. In contrast, IL-8 receptor mRNA was only detected in stromal cells. The expression of IL-8 in GLCs and KGN cells was increased by addition of IL-1beta and TNFalpha. Interleukin-8 increased the proliferation of ovarian stromal cells. The expression of pIkappaB in KGN cells was induced by IL-1beta, and the effects were reduced by APDC. CONCLUSION(S): Interleukin 8 induced by IL-1beta via activation of NF-kappaB in granulosa cells may have a role in the periovulatory period of follicular maturation.

Induction of hepatocyte growth factor in stromal cells by tumor-derived basic fibroblast growth factor enhances growth and invasion of endometrial cancer.

Tumor progression is often regulated through interactions between carcinoma cells and host stromal cells. In this study of endometrial cancer, we investigated one mechanism potentially involved in hepatocyte growth factor (HGF)-mediated cancer-stromal interactions. Endometrial cancer cells (HEC-1 and ISHIKAWA) expressed the c-met receptor, but HGF did not. HGF, however, did stimulate the proliferation and invasion of these cells. The HGF gene was expressed in stromal cells, which had been separated from primary cultures of endometrial cancers, 6.4 times more than in isolated normal endometrial stromal cells. Immunohistochemical staining revealed immunoreactive HGF in cancer stromal cells, the staining intensity being more pronounced in cancer tissue than in normal endometrium. The conditioned medium from normal epithelial cells and cancer cell lines induced HGF production in normal stromal cells. We identified basic fibroblast growth factor as an HGF inducer derived from endometrial cancer cell lines. Basic fibroblast growth factor derived from tumor cells may induce HGF in endometrial stromal cells, whereas stromal cell-derived HGF leads to the invasive growth of carcinoma cells. These interactions, mediated by HGF and HGF inducers, may play a significant role in the progression of endometrial cancer.