Reversal of silver sulfadiazine-impaired wound healing by epidermal growth factor

Silver sulfadiazine (Ag-SD) is a useful antibacterial agent for wound treatment. However, recent findings indicate that the compound delays the wound-healing process. That delay may be reversed by treatment with growth factors. The purpose of this study, was to evaluate the cyto-protective effect of epidermal growth factor (EGF) against Ag-SD treated keratinocytes and to investigate the reversibility of the impaired wound-healing process by the co-supplementation of EGF. Four types of drug-loaded collagen sponge dressings with different concentrations of Ag-SD, EGF and Ag-SD + EGF were prepared. An immortalized keratinocyte, HaCaT cells, were cultured in 35-mm Petri-dish using Dulbecco's Modified Eagle's Minimal Essential Medium (DMEM) with 10% FBS. Cultures were treated with the samples submerged, and viabilities of cultures were evaluated using MTT assay. The wound heal efficacy was evaluated in a partial thickness burn mouse model. Cells treated with EGF showed a cyto-protective effect on 1% Ag-SD treated cells with significant increase in viable cell numbers at concentrations ranging from 1 to 50 microg/ml. The cytotoxicity of Ag-SD impaired wound healing, while the addition of EGF could reverse the impairment. This evidence suggests that EGF is a useful agent in the retardation of wound healing caused by Ag-SD. Therefore, a drug delivery system containing both EGF and Ag-SD, such as that used in the study, may be clinically relevant.     

Cytotoxic constituents from the roots ofAnthriscus sylvestris

Activity-guided fractionation of the roots of Anthriscus sylvestris resulted in the isolation and characterization of five cytotoxic compounds, deoxypodophyllotoxin (1), falcarindiol (2), and angeloyl podophyllotoxin (5) from the hexane soluble fraction and morelensin (3), bursehernin (4) from the chloroform soluble fraction. It is the first report of the occurrence of compound 5 in nature.     

A Novel Nicotinamide Adenine Dinucleotide Correction Method for Mitochondrial Ca2+ Measurement with FURA-2-FF in Single Permeabilized Ventricular Myocytes of Rat

Fura-2 analogs are ratiometric fluoroprobes that are widely used for the quantitative measurement of [Ca²⁺]. However, the dye usage is intrinsically limited, as the dyes require ultraviolet (UV) excitation, which can also generate great interference, mainly from nicotinamide adenine dinucleotide (NADH) autofluorescence. Specifically, this limitation causes serious problems for the quantitative measurement of mitochondrial [Ca²⁺], as no available ratiometric dyes are excited in the visible range. Thus, NADH interference cannot be avoided during quantitative measurement of [Ca²⁺] because the majority of NADH is located in the mitochondria. The emission intensity ratio of two different excitation wavelengths must be constant when the fluorescent dye concentration is the same. In accordance with this principle, we developed a novel online method that corrected NADH and Fura-2-FF interference. We simultaneously measured multiple parameters, including NADH, [Ca²⁺], and pH/mitochondrial membrane potential; Fura-2-FF for mitochondrial [Ca²⁺] and TMRE for Ψ_m or carboxy-SNARF-1 for pH were used. With this novel method, we found that the resting mitochondrial [Ca²⁺] concentration was 1.03 µM. This 1 µM cytosolic Ca²⁺ could theoretically increase to more than 100 mM in mitochondria. However, the mitochondrial [Ca²⁺] increase was limited to ~30 µM in the presence of 1 µM cytosolic Ca²⁺. Our method solved the problem of NADH signal contamination during the use of Fura-2 analogs, and therefore the method may be useful when NADH interference is expected.     

Melatonin suppresses NO-induced apoptosis via induction of Bcl-2 expression in PGT-beta immortalized pineal cells

In the present study, we investigated whether melatonin would prevent nitric oxide (NO)-induced apoptotic death of PGT-beta immortalized pineal cells. To examine the protective effect of melatonin, cytotoxicity assay, DNA fragmentation analysis, caspase-3 activity assay, and Western blotting for caspase-3 and poly(ADP-ribose) polymerase (PARP) were performed. Treatment of cells with S-nitroso-N-acetylpenicillamine (SNAP), an NO donor, was shown to induce apoptotic cell death in a dose-dependent manner, and pretreatment with melatonin (0.1 mm) attenuated the occurrence of NO-induced apoptotic cell death. DNA fragmentation in response to NO was also arrested by melatonin. Caspase-3 activity induced by NO was decreased with melatonin treatment. Furthermore, the active fragments of caspase-3 and PARP were almost completely absent following exposure to melatonin. To elucidate the protective mechanisms of action of melatonin, Western blot analyses for Bcl-2 expression and cytochrome c release were carried out. Pretreatment with melatonin (0.1 mm) induced the expression of Bcl-2 and suppressed the release of cytochrome c into the cytosol, thereby arresting NO-induced apoptotic cell death. These results suggest that the antiapoptotic effect of melatonin is associated with induction of Bcl-2 expression in PGT-beta cells, which in turn blocks caspase-3 activation and inhibits cytochrome c release into the cytosol.     

Factors associated with Successful Responses to Ganglion Impar Block: A Retrospective Study

Background: The ganglion impar (ganglion of Walther) block has been used to manage coccygeal and perineal (perianal and genital) pain due to both benign and malignant causes. However, the factors associated with successful responses to ganglion impar block are unknown. Therefore, in the present study, we aimed to identify the independent factors associated with successful responses to ganglion impar block in patients with chronic pain in coccygeal and perineal regions.Methods: From January 2008 to December 2017, we performed a retrospective review of 106 patients who underwent ganglion impar block. Patients were considered successful responders if they reported a decrease of more than 50% or 4 points on the 11-point (0 = no pain and 10 = worst possible pain) numerical rating scale 1 month after the procedure, while others were considered non-responders. Logistic regression analysis was performed to identify factors independently associated with successful responses at 1 month after the procedure.Results: Multivariable logistic regression analysis showed that cancer-related causes were significantly associated with successful responses at 1 month after ganglion impar block (odds ratio = 2.60, 95% confidence interval = 1.05 to 6.43, P = 0.038).Conclusion: Ganglion impar block may be more effective in cancer-related pain than pain due to benign causes.     

Sixty-five gene-based risk score classifier predicts overall survival in hepatocellular carcinoma

Clinical application of the prognostic gene expression signature has been delayed due to the large number of genes and complexity of prediction algorithms. In the current study we aimed to develop an easy-to-use risk score with a limited number of genes that can robustly predict prognosis of patients with hepatocellular carcinoma (HCC). The risk score was developed using Cox coefficient values of 65 genes in the training set (n = 139) and its robustness was validated in test sets (n = 292). The risk score was a highly significant predictor of overall survival (OS) in the first test cohort (P = 5.6 × 10(-5), n = 100) and the second test cohort (P = 5.0 × 10(-5) , n = 192). In multivariate analysis, the risk score was a significant risk factor among clinical variables examined together (hazard ratio [HR], 1.36; 95% confidence interval [CI], 1.13-1.64; P = 0.001 for OS). CONCLUSION: The risk score classifier we have developed can identify two clinically distinct HCC subtypes at early and late stages of the disease in a simple and highly reproducible manner across multiple datasets.     

Bilateral hyperexcitability of thalamic VPL neurons following unilateral spinal injury in rats

In the present study, we have examined whether spinal hemisection injury induces changes in the electrophysiological properties of thalamic ventral posteriorlateral (VPL) neurons in rats. Male Sprague–Dawley rats were subjected to unilateral spinal cord injury by transverse hemisection at the T13 spinal segment. Four weeks after the T13 spinal hemisection, the injured rats displayed robust allodynic behaviors on both sides of hindpaws compared to sham controls (P < 0.05). Extracellular recordings taken 4 weeks after the hemisection revealed that wide dynamic range (WDR) neurons had significantly increased spontaneous and brush-, pressure-, and pinch-evoked activities, respectively, on both sides of the thalamic VPL regions (P < 0.05). In contrast, low threshold (LT) neurons showed only an increase in the brush-evoked activity compared to sham controls (P < 0.05). However, afterdischarge activity in both types of neurons showed no changes. In addition, both sides of the thalamic VPL regions showed higher incidences of WDR neurons. In conclusion, our data demonstrate that spinal unilateral injury induces bilaterally increased evoked activity in thalamic VPL neurons.