Mitochondrial dynamics regulate melanogenesis through proteasomal degradation of MITF via ROS‐ERK activation

Mitochondrial dynamics control mitochondrial functions as well as their morphology. However, the role of mitochondrial dynamics in melanogenesis is largely unknown. Here, we show that mitochondrial dynamics regulate melanogenesis by modulating the ROS‐ERK signaling pathway. Genetic and chemical inhibition of Drp1, a mitochondrial fission protein, increased melanin production and mitochondrial elongation in melanocytes and melanoma cells. In contrast, down‐regulation of OPA1, a mitochondria fusion regulator, suppressed melanogensis but induced massive mitochondrial fragmentation in hyperpigmented cells. Consistently, treatment with CCCP, a mitochondrial fission chemical inducer, also efficiently repressed melanogenesis. Furthermore, we found that ROS production and ERK phosphorylation were increased in cells with fragmented mitochondria. And inhibition of ROS or ERK suppressed the antimelanogenic effect of mitochondrial fission in α‐MSH‐treated cells. In addition, the activation of ROS‐ERK pathway by mitochondrial fission induced phosphorylation of serine73 on MITF accelerating its proteasomal degradation. In conclusion, mitochondrial dynamics may regulate melanogenesis by modulating ROS‐ERK signaling pathway.     

Synthetic chitinase gene driven by root-specific LjNRT2 and AtNRT2.1 promoters confers resistance to Fusarium oxysporum in transgenic tobacco and tomato

Fusarium wilt is a soil-borne disease causing substantial yield losses in various crops and vegetables. We have previously reported the synthetic chitinase (NIC) gene (1.2 kb), in which codon usage of fungus, replaced with that of plant, conferred resistance against Botrytis cinerea. In this study, the NIC or GUS gene was linked to two root-specific promoters, LjNRT2 or AtNRT2.1 (nitrate transporter 2), derived from Lotus japonica and Arabidopsis thaliana, respectively. Transgenic tobacco lines expressing LjNRT2-GUS and LjNRT2-NIC, and tomato lines expressing AtNRT2.1-NIC, were produced by Agrobacterium-mediated transformation. GUS histochemical staining was observed in vascular regions of the roots but was conspicuously absent in the leaves of transgenic plants. Western blot analysis showed the production of NIC proteins in the roots but not in the leaves of transgenic tobacco and tomato lines. These results indicate that LjNRT2 and AtNRT2.1 promoters expressed transgenes in a root-specific manner. When in vitro whole plant resistance assay against Fusarium oxysporum was conducted, transgenic plants showed increased levels of resistance compared to non-transgenic plants. Antifungal activities of the root extract against spore germination of F. oxysporum showed lower CFU (colony-forming unit) than those of the leaf extract. Root colonization assay against F. oxysporum showed much lower CFU values in the roots of transgenic plants than in those of non-transgenic plants. These results suggest that NIC gene triggered by the root-specific promoters successfully expressed only in the roots and conferred increased levels of resistance against the root pathogen, F. oxysporum.     

Body Mass Index and Mortality in Korean Intensive Care Units: A Prospective Multicenter Cohort Study

Background

The level of body mass index (BMI) that is associated with the lowest mortality in critically ill patients in Asian populations is uncertain. We aimed to examine the association of BMI with hospital mortality in critically ill patients in Korea.

Methods

We conducted a prospective multicenter cohort study of 3,655 critically ill patients in 22 intensive care units (ICUs) in Korea. BMI was categorized into five groups: <18.5, 18.5 to 22.9, 23.0 to 24.9 (the reference category), 25.0 to 29.9, and ≥30.0 kg/m².

Results

The median BMI was 22.6 (IQR 20.3 to 25.1). The percentages of patients with BMI<18.5, 18.5 to 22.9, 23.0 to 24.9, 25.0 to 29.9, and ≥30.0 were 12, 42.3, 19.9, 22.4, and 3.3%, respectively. The Cox-proportional hazard ratios with exact partial likelihood to handle tied failures for hospital mortality comparing the BMI categories <18.5, 18.5 to 22.9, 25.0 to 29.9, and ≥30.0 with the reference category were 1.13 (0.88 to 1.44), 1.03 (0.84 to 1.26), 0.96 (0.76 to 1.22), and 0.68 (0.43 to 1.08), respectively, with a highly significant test for trend (p = 0.02).

Conclusions

A graded inverse association between BMI and hospital mortality with a strong significant trend was found in critically ill patients in Korea.     

TLR4-Mediated Expression of Mac-1 in Monocytes Plays a Pivotal Role in Monocyte Adhesion to Vascular Endothelium

Toll-like receptor 4 (TLR4) is known to mediate monocyte adhesion to endothelial cells, however, its role on the expression of monocyte adhesion molecules is unclear. In the present study, we investigated the role of TLR4 on the expression of monocyte adhesion molecules, and determined the functional role of TLR4-induced adhesion molecules on monocyte adhesion to endothelial cells. When THP-1 monocytes were stimulated with Kdo2-Lipid A (KLA), a specific TLR4 agonist, Mac-1 expression was markedly increased in association with an increased adhesion of monocytes to endothelial cells. These were attenuated by anti-Mac-1 antibody, suggesting a functional role of TLR4-induced Mac-1 on monocyte adhesion to endothelial cells. In monocytes treated with MK886, a 5-lipoxygenase (LO) inhibitor, both Mac-1 expression and monocyte adhesion to endothelial cells induced by KLA were markedly attenuated. Moreover, KLA increased the expression of mRNA and protein of 5-LO, suggesting a pivotal role of 5-LO on these processes. In in vivo studies, KLA increased monocyte adhesion to aortic endothelium of wild-type (WT) mice, which was attenuated in WT mice treated with anti-Mac-1 antibody as well as in TLR4-deficient mice. Taken together, TLR4-mediated expression of Mac-1 in monocytes plays a pivotal role on monocyte adhesion to vascular endothelium, leading to increased foam cell formation in the development of atherosclerosis.     

Stereotactic Body Radiation Therapy as an Alternative Treatment for Small Hepatocellular Carcinoma

Background

Even with early stage hepatocellular carcinoma (HCC), patients are often ineligible for surgical resection, transplantation, or local ablation due to advanced cirrhosis, donor shortage, or difficult location. Stereotactic body radiation therapy (SBRT) has been established as a standard treatment option for patients with stage I lung cancer, who are not eligible for surgery, and may be a promising alternative treatment for patients with small HCC who are not eligible for curative treatment.

Materials and Methods

A registry database of 93 patients who were treated with SBRT for HCC between 2007 and 2009 was analyzed. A dose of 10-20 Gy per fraction was given over 3-4 consecutive days, resulting in a total dose of 30-60 Gy. The tumor response was determined using dynamic computed tomography or magnetic resonance imaging, which was performed 3 months after completion of SBRT.

Results

The median follow-up period was 25.6 months. Median size of tumors was 2 cm (range: 1-6 cm). Overall patients’ survival rates at 1 and 3 years were 86.0% and 53.8%, respectively. Complete and partial tumor response were achieved in 15.5% and 45.7% of patients, respectively. Local recurrence-free survival rate was 92.1% at 3 years. Most local failures were found in patients with HCCs > 3 cm, and local control rate at 3 years was 76.3% in patients with HCC > 3 cm, 93.3% in patients with tumors between 2.1-3 cm, and 100% in patients with tumors ≤ 2 cm, respectively. Out-of-field intrahepatic recurrence-free survival rates at 1 and 3 years were 51.9% and 32.4%, respectively. Grade ≥ 3 hepatic toxicity was observed in 6 (6.5%).

Conclusions

SBRT was effective in local control of small HCC. SBRT may be a promising alternative treatment for patients with small HCC which is unsuitable for other curative therapy.     

Production of offspring from cloned transgenic RFP female dogs and stable generational transmission of the RFP gene

The purpose of this study was to analyze the reproductive ability of transgenic female dogs born bysomatic cell nuclear transfer and to determine inheritance of the red fluorescent protein (RFP) transgene. The four founder transgenic bitches (F0) reached puberty at 340.8 ± 39.6 days after birth and were bred with wild-type male dogs by natural mating or by artificial insemination. The bitches all became pregnant and successfully delivered 13 puppies (F1), of which two females were bred with wild-type dogs to deliver 7 offspring (F2), including 1 stillbirth. Among the 19 live offspring, 10 puppies showed emission of RFP under UV light and the presence of the RFP transgene was confirmed by genomic PCR and Southern blot analyses. In conclusion, transgenic RFP female dogs exhibited normal reproductive ability and expression of the transgene was demonstrated in F1 and F2 generations.     

Poly(carboxybetaine methacrylamide)-modified nanoparticles: a model system for studying the effect of chain chemistry on film properties, adsorbed protein conformation, and clot formation kinetics

Nonfouling polymer architectures are considered important to the successful implementation of many biomaterials. It is thought that how these polymers induce conformational changes in proteins upon adsorption may dictate the fate of the device being utilized. Herein, oxidized silicon nanoparticles (SiNP) were modified with various forms of poly(carboxybetaine methacrylamide) (PCBMA) for the express purpose of understanding how polymer chemistry affects film hydration, adsorbed protein conformation, and clot formation kinetics. To this end, carboxybetaine monomers differing in intercharge separating spacer groups were synthesized, and nitroxide-mediated free radical polymerization (NMP) was conducted using alkoxyamine initiators with hydrophobic (TEMPO) and hydrophilic (β-phosphonate) terminal groups. The physical properties (surface composition, thickness, grafting density, etc.) of the resulting polymer-SiNP conjugates were quantified using several techniques, including Fourier transform infrared (FTIR) spectroscopy, dynamic light scattering (DLS), and thermogravimetric analysis (TGA). The effect of spacer group on the surface charge density was determined using zeta potential measurements. Three proteins, viz., lysozyme, bovine α-lactalbumin, and human serum albumin, were used to evaluate the effect film properties (charge, hydration, end-group) have on adsorbed protein conformation, as determined by circular dichroism (CD), fluorescence spectroscopy, and fluorescence quenching techniques. Hemocompatibility of these surfaces was observed by measuring clot formation kinetics using the plasma recalcification time assay. It was found that chain chemistry, as opposed to end-group chemistry, was a major determiner for water structure, adsorbed protein conformation, and clotting kinetics. It is thought that the systematic evaluation of how both chain (internal) and end-group (external) polymer properties affect film hydration, protein conformation, and clot formation will provide valuable insight that can be applied to all engineered surfaces for biomedical applications.     

Photo‐Carrier Recombination in Polymer Solar Cells Based on P3HT and Silole‐Based Copolymer

Photo‐current loss in donor‐acceptor (DA) polymer‐fullerene bulk heterojunction solar cells was studied via carrier transport and recombination measurements. Focusing on the DA polymer poly((4,4‐dioctyldithieno (3,2‐b:2',3'‐d) silole)‐2,6‐diyl‐alt‐(2,1,3‐benzothiadiazole)‐4,7‐diyl) (DTS‐BTD) we found that the carrier transport is well‐balanced and attribute the loss mechanism in DTS‐BTD solar cells to carrier recombination. Using carrier extraction with linear increasing voltage (photo‐CELIV) and transient photo‐voltage (TPV), we show that carrier recombination plays an important role in photo‐current extraction at open circuit conditions due to increase in photo‐excited carrier concentration. Delay time dependent photo‐CELIV and temperature dependent transport studies suggest that the recombination rate is related to the degree of energetic disorder in the polymer: fullerene blends.