Galuteolin, identified in the extract of thymus quinquecostatus flowers, is involved in inhibiting melanin biosynthesis in B16/F10 melanoma cells.

To enhance the skin whitening effect, tyrosinase activity and melanin biosynthesis needs to be suppressed in the skin. To achieve this goal, we examined the extract of Thymus quinquecostatus flowers, and identified a functional ingredient, galuteolin. Galuteolin effectively inhibited melanin biosynthesis in B16/F10 cells, partially suppressing tyrosinase activity. Therefore, this study suggests that galuteolin can be used as a cosmetic ingredient for skin whitening.

Flexible electrochemical glucose biosensor based on GOx/gold/MoS2/gold nanofilm on the polymer electrode.

The need for flexible biosensors has increased because of their potential applications for point-of-care diagnosis and wearable biosensors. However, flexible biosensors have low sensitivity due to the flexibility of the electrode, and their fabrication involves complex processes. To overcome these limitations, a flexible electrochemical enzyme biosensor was developed in this study by immobilizing an enzyme on the flexible polymer electrode modified with a gold/MoS2/gold nanofilm. The fabrication process involved sputter deposition of gold, spin coating of MoS2, and sputter deposition of gold on the flexible polymer electrode (commercially available Kapton® polyimide film). The flexible glucose biosensor was made by immobilization of glucose oxidase on a flexible electrode by using a chemical linker. The detection limit for glucose was estimated to be 10 nM, which indicates more sensitivity as compared with a previously reported flexible glucose sensor. This sensitivity is due to the facilitation of electron transfer by MoS2. The flexure extension of this biosensor was estimated at 3.48 mm, which is much higher than that of the rigid sensor using a gold-coated silicon electrode (0.09 mm), according to measurements with a micro-fatigue tester. The proposed flexible biosensor composed of the enzyme/gold/MoS2/gold nanofilm on the polymer electrode can be used as a flexible sensing platform for developing wearable biosensing systems because of its high sensitivity, high flexibility, and simple fabrication process.

Effect of angiotensin II type 1 receptor blocker and angiotensin converting enzyme inhibitor on the intraocular growth factors and their receptors in streptozotocin-induced diabetic rats.

AIM: To investigate the effect of angiotensin II type 1 receptor blocker (ARB) and angiotensin converting enzyme inhibitor (ACEI) on intraocular growth factors and their receptors in streptozotocin-induced diabetic rats. METHODS: Forty Sprague-Dawley rats were divided into 4 groups: control, diabetes mellitus (DM), candesartan-treated DM, and enalapril-treated DM (each group, n=10). After the induction of DM by streptozotocin, candesartan [ARB, 5 mg/(kg·d)] and enalapril [ACEI, 10 mg/(kg·d)] were administered to rats orally for 4wk. Vascular endothelial growth factor (VEGF) and angiotensin II (Ang II) concentrations in the vitreous were measured using enzyme-linked immunosorbent assays, and VEGF receptor 2 and angiotensin II type 1 receptor (AT1R) levels were assessed at week 4 by Western blotting. RESULTS: Vitreous Ang II levels were significantly higher in the DM group and candesartan-treated DM group than in the control (P=0.04 and 0.005, respectively). Vitreous AT1R increased significantly in DM compared to the other three groups (P<0.007). Candesartan-treated DM rats showed higher vitreal AT1R concentration than the enalapril-treated DM group and control (P<0.001 and P=0.005, respectively). No difference in vitreous Ang II and AT1R concentration was found between the enalapril-treated DM group and control. VEGF and its receptor were below the minimum detection limit in all 4 groups. CONCLUSION: Increased Ang II and AT1R in the hyperglycemic state indicate activated the intraocular renin-angiotensin system, which is inhibited more effectively by systemic ACEI than systemic ARB.

Choroidal Watershed Zone and Growth of Polypoidal Choroidal Vasculopathy.

PURPOSE: To investigate the topographic relationship between polypoidal choroidal vasculopathy (PCV) growth and choroidal watershed zones (CWZ) by using indocyanine green angiography (ICGA). MATERIALS AND METHODS: We evaluated PCV lesions smaller than the CWZ at baseline and followed up more than 6 months. The CWZ was traced in the early phase ICGA at baseline. The vascular lesion of PCV was traced in baseline and follow-up ICGA. These traces were overlapped and topographic relationships between CWZs and PCV growth were evaluated. RESULTS: Among 31 eyes of 31 patients, enlargement of a PCV lesion was observed in 20 patients (64.5%) at mean follow-up of 30.4 months (6-68 months). A topographical relationship between the CWZ and PCV growth was demonstrated in 14 eyes (70.0%), as the shape of the PCV lesion conformed to the boundary of the CWZ, and/or the growth of the branching vascular network was aligned to the direction of the CWZ extension to the periphery. Growth beyond the boundary of the CWZ was noted in 9 eyes (45.0%), however growth area was greater inside the CWZ than outside in all eyes. Of 15 eyes of extrafoveal lesion at baseline, 8 eyes in which the vascular lesion progressed to the fovea had the CWZ involving the fovea, whereas PCV in three eyes with an extrafoveal CWZ remained as a non-subfoveal disease after mean follow-up of 17.0 months (p = 0.019). CONCLUSIONS: A subfoveal CWZ was related to PCV growth to the fovea. Topographical relationships between PCV growth and the CWZ suggest that choroidal circulation is a predisposition for PCV growth.

Topographical relationship between the choroidal watershed zone and submacular idiopathic choroidal neovascularisation.

AIMS: To investigate the relationship between idiopathic choroidal neovascularisation (CNV) and choroidal watershed zones (CWZs) using indocyanine green angiography (ICGA). DESIGN: Multicentre, retrospective, interventional case series. METHODS: The medical records and ICGA findings of 44 patients (44 eyes) diagnosed with idiopathic CNV were reviewed. CWZs, defined as hypofluorescence that disappeared during the early phase of ICGA, were classified, and the findings were compared with those of a control group of 30 eyes. The topographical relationship between CWZs and CNV was evaluated. Visual acuity and recurrence were analysed according to the CWZ classification. RESULTS: The CNV lesion was subfoveal in 16 eyes, juxtafoveal in 12 eyes and extrafoveal in 16 eyes. The most common types of CWZs were stellate (23 eyes, 52.3%) and vertical (19 eyes, 43.2%). CWZs involving the fovea were seen in more patients with idiopathic CNV (37 eyes, 84.1%) than in the control group (11 eyes, 36.7%, p<0.001). The topographical relationship between CWZs and CNV was determined in 42 eyes (95.5%), with the CNV located within the CWZ in 39 eyes and at the margin in 3 eyes. Extrafoveal CNV was within the CWZ in all 16 affected eyes. At 6 months, visual acuity was significantly worse in patients with subfoveal CNV (p=0.028) or stellate CWZs (p=0.039). CONCLUSIONS: The findings of a CWZ were related to the location and functional outcome of idiopathic CNV. Our results suggest that choroidal circulation is a predisposing factor for the development of CNV in young patients.

Evaluation of Macular Function Using Pattern Electroretinogram in Idiopathic Epiretinal Membrane.

PURPOSE: Pattern electroretinogram (PERG) in idiopathic epiretinal membrane (ERM) was investigated to find the correlation of the ganglion cell function with postoperative visual acuity. DESIGN: This was a retrospective consecutive chart review. METHODS: Medical records of 24 eyes that underwent vitrectomy and membrane peeling for idiopathic ERM were reviewed retrospectively. The amplitude and implicit time of P50 and N95 in preoperative PERG were analyzed to find correlation with visual acuity and foveal thickness. The ratio of the parameters in involved eyes to those in healthy fellow eyes was calculated for analysis. RESULTS: Visual acuity (logarithm of the minimum angle of resolution) improved from 0.53 at baseline to 0.34 at 6 months (P = 0.003). Foveal thickness decreased significantly from 488.3 µm at baseline to 374.7 µm (P = 0.001). The preoperative N95 amplitude ratio was significantly correlated with visual acuity at 6 months after ERM removal (r = -0.423, P = 0.040), whereas the amplitude of P50 and implicit time of both waves showed no significant correlation with postoperative visual acuity. The implicit time ratio of P50 (r = 0.530, P = 0.008) and N95 (r = 0.436, P = 0.033) showed significant correlation with preoperative foveal thickness on optical coherence tomography. CONCLUSIONS: N95 amplitude in PERG was a predictor of visual outcomes after ERM surgery. These results suggest the correlation of postoperative visual acuity with the function of the ganglion cell layer, which is the closest cell layer to be affected by ERM.

Autologous advanced tenon grafting combined with conjunctival flap in scleromalacia after pterygium excision.

Purpose. To evaluate the efficacy of autologous tenon grafting combined with conjunctival flap as a treatment for scleromalacia or scleral thinning after pterygium excision without any additional donor graft tissue. Methods. Twenty-six cases underwent autologous advanced tenon grafting combined with sliding or rotating conjunctival flap for scleromalacia after pterygium surgery ranging from 2 years to 30 years. The extent of scleral defect measured from 2.0 mm to 6.8 mm in diameter. The cosmetic outcome was defined as complete resolution of scleromalacia or completely conjunctival reepithelialization and firm adhesion between subtenon and scleral tissue over scleral thinning without significant complications. Results. All cases achieved the covering of conjunctival and tenon or subtenon tissue over scleromalacia or scleral thinning with this procedure. Preoperative pain, inflammation, and choroidal exposure disappeared after surgery. Immediate postoperative complications, such as large wound dehiscence or reopening of the scleral wound, did not occur in any of the patients. There were no significant clinical complications during the mean postoperative follow-up period of 14.17 months in all cases. Conclusions. We obtained excellent outcome with fewer complications after autologous advanced tenon graft and conjunctival flap, without an additional donor graft, in scleromalacia or scleral thinning caused by previous pterygium excision.

Reduced-fluence photodynamic therapy in polypoidal choroidal vasculopathy nonresponsive to ranibizumab.

BACKGROUND AND OBJECTIVE: To evaluate the effect of reduced-fluence photodynamic therapy (PDT) on polypoidal choroidal vasculopathy (PCV) unresponsive to intravitreal ranibizumab. PATIENTS AND METHODS: Patients with PCV unresponsive to ranibizumab administered 3 months consecutively who then received reduced-fluence PDT were retrospectively surveyed. Nonresponders were defined as patients having no reduction in intraretinal and/or subretinal fluid after 3 consecutive treatments. RESULTS: In total, 22 of 104 eyes (21.2%) were non-responders, and 16 of 22 nonresponders received reduced-fluence PDT. Nine eyes achieved complete fluid resolution, and six had reduced but persistent fluid. In one eye, fluid persisted at 6 months despite an additional anti-vascular endothelial growth factor (anti-VEGF) injection after reduced-fluence PDT. Mean macular thickness decreased significantly at 3 and 6 months after PDT, but the mean visual acuity was worse than baseline. CONCLUSION: Reduced-fluence PDT in nonresponders gradually decreased intraretinal and/or subretinal fluid over several months but did not maintain visual acuity.

Serous retinal detachment accompanied by MEWDS in a myopic patient with dome-shaped macula.

Macular serous retinal detachment (MSRD) is a rare complication in highly myopic patients with an inferior staphyloma, tilted disc, or dome-shaped macula. Multiple evanescent white dot syndrome (MEWDS) presents with sudden visual loss and multiple yellowish dots that resolve spontaneously within several weeks. The authors report the development and spontaneous resolution of subretinal fluid accompanied by MEWDS in a myopic patient with a dome-shaped macula. Dysfunction of the retinal pigment epithelium due to MEWDS likely induced temporary MSRD in this patient.

CD4+CD25+ regulatory T cell depletion modulates anxiety and depression-like behaviors in mice.

Stress has been shown to suppress immune function and increase susceptibility to inflammatory disease and psychiatric disease. CD4(+)CD25(+) regulatory T (Treg) cells are prominent in immune regulation. This study was conducted to determine if anti-CD25 antibody (Ab) mediated depletion of Treg cells in mice susceptibility to stress-induced development of depression-like behaviors, as well as immunological and neurochemical activity. To accomplish this, an elevated plus-maze test (EPM), tail suspension test (TST), and forced swim test (FST) were used to examine depression-like behaviors upon chronic immobilization stress. Immune imbalance status was observed based on analysis of serum cytokines using a mouse cytometric bead array in conjunction with flow cytometry and changes in the levels of serotonin (5-HT) and dopamine (DA) in the brain were measured by high performance liquid chromatography (HPLC). The time spent in the open arms of the EPM decreased significantly and the immobility time in the FST increased significantly in the anti-CD25 Ab-treated group when compared with the non stressed wild-type group. In addition, interlukin-6 (IL-6), tumor necrosis factor-á (TNF-á), interlukin-2 (IL-2), interferon-gamma (IFN-γ), interlukin-4 (IL-4) and interlukin-17A (IL-17A) concentrations were significantly upregulated in the stressed anti-CD25 Ab-treated group when compared with the non stressed wild-type group. Furthermore, the non stressed anti-CD25 Ab-treated group displayed decreased 5-HT levels within the hippocampus when compared with the non stressed wild-type group. These results suggest that CD4(+)CD25(+) Treg cell depletion modulated alterations in depressive behavior, cytokine and monoaminergic activity. Therefore, controlling CD4(+)CD25(+) Treg cell function during stress may be a potent therapeutic strategy for the treatment of depression-like symptoms.

T-bet deficient mice exhibit resistance to stress-induced development of depression-like behaviors.

T-bet, a Th1-specific T-box transcription factor, regulates Th1 development by inducing endogenous Th1 cytokines and IFN-γ. This study was conducted to determine if T-bet knockout mice exhibit resistance to stress-induced development of depression-like behaviors. The T-bet knockout mice significantly reduced depressive-like behaviors provoked by repeated restraint stress in an elevated plus-maze test (EPM), tail suspension test (TST), and forced swim test (FST). Moreover, stress-induced elevations of the pro-inflammatory cytokines were attenuated in T-bet deficient group. These results suggest that T-bet directly mediated stress-induced depression. Therefore, understanding T-bet function during stress represents an additional treatment strategy for depression.

A Parametric Study on the Immunomodulatory Effects of Electroacupuncture in DNP-KLH Immunized Mice.

This study was conducted to compare the effects of low frequency electroacupuncture (EA) and high frequency EA at acupoint ST36 on the production of IgE and Th1/Th2 cytokines in BALB/c mice that had been immunized with 2,4-dinitrophenylated keyhole limpet protein (DNP-KLH), as well as to investigate the difference in the immunomodulatory effects exerted by EA stimulations at acupoint ST36 and at a non-acupoint (tail). Female BALB/c mice were divided into seven groups: normal (no treatments), IM (immunization only), ST36-PA (IM + plain acupuncture at ST36), ST36-LEA (IM + low frequency (1 Hz) EA at ST36), ST36-HEA (IM + high frequency (120 Hz) EA at ST36), NA-LEA (IM + low frequency (1 Hz) EA at non-acupoint) and NA-HEA (IM + high frequency (120 Hz) EA at non-acupoint). EA stimulation was performed daily for two weeks, and total IgE, DNP-KLH specific IgE, IL-4 and IFN-γ levels were measured at the end of the experiment. The results of this study showed that the IgE and IL-4 levels were significantly suppressed in the ST36-LEA and ST36-HEA groups, but not in the NA-LEA and NA-HEA groups. However, there was little difference in the immunomodulatory effects observed in the ST36-LEA and ST36-HEA groups. Taken together, these results suggest that EA stimulation-induced immunomodulation is not frequency dependent, but that it is acupoint specific.

Methyl gallate exhibits potent antitumor activities by inhibiting tumor infiltration of CD4+CD25+ regulatory T cells.

CD4(+)CD25(+) regulatory T (Treg) cells play crucial roles in the host response to tumors. Increasing evidence supports the existence of elevated numbers of Treg cells in solid tumors and hematologic malignancies. In this study, the effects of methyl gallate on Treg cells were examined. Methyl gallate inhibited Treg cell-suppressive effects on effector CD4(+) T cells and Treg migration toward tumor environment. The expression of Treg surface markers including CTLA-4, CCR4, CXCR4, and glucocorticoid-induced TNFR was significantly suppressed upon methyl gallate treatment. Furthermore, forkhead box P3 (Foxp3) expression was also significantly decreased by methyl gallate, suggesting that the suppressive effects of methyl gallate on Treg were medicated by decrease of Treg-specific transcription factor Foxp3. In tumor-bearing hosts, methyl gallate treatment substantially reduced tumor growth and prolonged the survival rate. In contrast, nu/nu mice did not show decreased tumor progression in response to methyl gallate. In addition, in tumor-bearing Treg-depleted mice, tumor growth and the survival rates were not changed by methyl gallate treatment, strongly suggesting that the main therapeutic target of methyl gallate in tumor suppression was related to modulation of the CD4(+)CD25(+) Treg cell functions. In the spleen of tumor-bearing mice, methyl gallate treatment induced a significant decrease in the CD4(+)CD25(+)Foxp3(high) Treg cell population. Especially, the number of tumor-infiltrating CD25(+)Foxp3(high) Treg cells was significantly lower in methyl gallate-treated mice. These results suggest that methyl gallate can be used to reverse immune suppression and as a potentially useful adjunct for enhancing the efficacy of immune-based cancer therapy.

CD4+CD25+ regulatory T cells attenuate cisplatin-induced nephrotoxicity in mice.

Nephrotoxicity limits the use of cisplatin, a widely used chemotherapeutic agent for treatment of various malignancies. Overall, CD4+ T cells mediate cisplatin-induced renal injury; however, the CD4+CD25+ regulatory T-cell subset (CD4+CD25+ Treg) has broad suppressive effects on many different cell types. In this study, we determined whether CD4+CD25+ Treg cells had protective effects against cisplatin-induced acute renal injury in nu/nu mice that lack mature T cells. In these mice, there was marked attenuation of the decreased survival, renal dysfunction and tubular injury, renal tumor necrosis factor-α, and interleukin-1ß cytokine levels. Furthermore, renal macrophage accumulation was reduced in CD4+CD25+ Treg cell-adoptive transferred nu/nu mice compared with control mice. Infusion of CD4+CD25+Treg cells into wild-type Balb/c mice reduced serum blood urea nitrogen and creatinine levels equivalent to those in nu/nu mice and extended their survival time after cisplatin injection. In contrast, depletion of CD4+CD25+ Treg cells in wild-type mice exacerbated kidney injury after cisplatin administration. Transcription factor Foxp3-positive cells (Treg cells) were detected in the kidneys of nu/nu mice after cisplatin injection. Our results suggest that CD4+CD25+ Treg cells directly affect cisplatin nephrotoxicity and their modulation represents an additional treatment strategy.

Microarray analysis of gene expression profiles in response to treatment with bee venom in lipopolysaccharide activated RAW 264.7 cells.

AIM OF THE STUDY: The therapeutic application of bee venom (BV) has been used in traditional medicine to treat diseases such as arthritis, rheumatism and pain. Macrophages produce molecules that are known to play roles in inflammatory responses. MATERIAL AND METHODS: We performed microarray analysis to evaluate the global gene expression profiles of RAW264.7 macrophage cells treated with BV. In addition, six genes were subjected to real-time PCR to confirm the results of the microarray. The cells were treated with lipopolysaccharide (LPS) or BV plus LPS for 30 min or 1h. RESULTS: 124 genes were found to be up-regulated and 158 were found to be down-regulated in cells that were treated with BV plus LPS for 30 min, whereas 211 genes were up-regulated and 129 were down-regulated in cells that were treated with BV plus LPS for 1h when compared with cells that were treated with LPS alone. Furthermore, the results of real-time PCR were similar to those of the microarray. BV inhibited the expression of specific inflammatory genes that were up-regulated by nuclear factor-kappa B in the presence of LPS, including mitogen-activated protein kinase kinase kinase 8 (MAP3K8), TNF, TNF-alpha-induced protein 3 (TNFAIP3), suppressor of cytokine signaling 3 (SOCS3), TNF receptor-associated factor 1 (TRAF1), JUN, and CREB binding protein (CBP). CONCLUSIONS: These results demonstrate the potent activity of BV as a modulator of the LPS-mediated nuclear factor-kappaB (NF-kappaB)/MAPK pathway in activated macrophages. In addition, these results can be used to understand other effects of BV treatment.

Molecular changes in remote tissues induced by electro-acupuncture stimulation at acupoint ST36.

To investigate the effects of electro-acupuncture (EA) treatment on regions remote from the application, we measured cellular, enzymatic, and transcriptional activities in various internal tissues of healthy rats. The EA was applied to the well-identified acupoint ST36 of the leg. After application, we measured the activity of natural killer cells in the spleen, gene expression in the hypothalamus, and the activities of antioxidative enzymes in the hypothalamus, liver and red blood cells. The EA treatment increased natural killer cell activity in the spleen by approximately 44%. It also induced genes related to pain, including 5-Hydroxytryptamine (serotonin) receptor 3a (Htr3a) and Endothelin receptor type B (Ednrb) in the hypothalamus, and increased the activity of superoxide dismutase in the hypothalamus, liver, and red blood cells. These findings indicate that EA mediates its effects through changes in cellular activity, gene expression, and enzymatic activity in multiple remote tissues. The sum of these alterations may explain the beneficial effects of EA.

The endogenous CCK mediation of electroacupuncture stimulation-induced satiety in rats.

A major satiety hormone, cholecystokinin (CCK) is well known to be released by electroacupuncture (EA) stimulation at certain body sites which elicits profound psychophysiological responses. Previous clinical and animal studies have shown that EA stimulation reduces food intake and body weight in both normal and obese subjects. The aim of the present study was to elucidate the satiety effect of EA stimulation and its mechanism related to CCK in rats. Here we show that EA stimulation at "Zusanli" (ST36) acupoint significantly reduced 30-min and 60-min food intake in 48-h fasted Sprague-Dawley rats, and such effect was reversed by a lorglumide (CCK-1 receptor antagonist, 10mg/kg, i.p.) pretreatment. The ST36 EA stimulation-induced satiety was not observed in CCK-1 receptor knockout, Otsuka Long-Evans Tokushima Fatty rats, but in their controls, Long-Evans Tokushima Otsuka rats. Subdiaphragmatic vagotomy also blocked the satiety effect of ST36 EA stimulation in Sprague-Dawley rats. These results suggest that ST36 EA stimulation elicits satiety in rats and this is mediated by the endogenous CCK signaling pathway.