Antimicrobial effect and mechanism of bovine lactoferrin against the potato common scab pathogen Streptomyces scabiei.

Lactoferrin (LF) is a multifunctional protein with a broad spectrum of antimicrobial activities. In this study, we investigated the antimicrobial activity of LF against the potato common scab pathogen Streptomyces scabiei, which causes severe damage to potato tubers. LF derived from bovine (bLF) had much higher activity against S. scabiei than human LF. The minimal inhibitory concentration of bLF was 3.9 µM. The effects of both apo-bLF (iron-free) and holo-bLF (iron-saturated) on S. scabiei were not different. Bovine lactoferricin (LFcinB), a short peptide with a length of 25 amino acid residues located in the N-terminal region of bLF, showed antimicrobial activity against S. scabiei, similar to that of bLF. These results indicated that the antimicrobial activity of bLF against S. scabiei cannot be attributed to its iron-chelating effect but to the bioactivity of its peptides. When S. scabiei was treated with the fusion protein of mCherry-LFcinB (red fluorescent protein) expressed in Escherichia coli, the pseudohyphal cells instantly glowed, indicating that the peptide electrostatically binds to the surface of S. scabiei. An assay of synthetic peptides, with modified number of arginine (Arg) and tryptophan (Trp) residues based on the antimicrobial center (RRWQWR) of LFcinB showed that Trp residues are implicated in the antimicrobial activity against S. scabiei; however, Arg residues are also necessary to carry Trp residues to the cell surface to fully exert its activity. Although the single amino acid effect of Trp had low activity, Trp derivatives showed much higher activity against S. scabiei, suggesting that the derivatives effectively bind to the cell surface (cell membrane) by themselves without a carrier. Thus, amino acid derivatives might be considered effective and alternative antimicrobial substances.

A Rare Case of Submassive Pulmonary Embolism with a Right Aberrant Subclavian Artery and Thrombosed Kommerell Diverticulum.

An 81-year-old man presented with shortness of breath and was referred to our hospital with suspected acute pulmonary embolism. Enhanced computed tomography revealed a right aberrant subclavian artery with a thrombosed Kommerell diverticulum (KD), as well as deep vein thrombosis in the left leg and bilateral pulmonary artery thrombosis. Thrombosis in the KD disappeared after one month of anticoagulation treatment with rivaroxaban. Thrombosis of a KD is a rare condition that may cause distal emboli and subclavian steal syndrome, although this syndrome was not present in this case. Rivaroxaban is an effective anticoagulant for treating thrombosis of a KD.

Impact of diesel exhaust exposure on the liver of mice fed on omega-3 polyunsaturated fatty acids-deficient diet.

Exposure to diesel exhaust (DE) exacerbates non-alcoholic fatty liver disease, and may systemically affect lipid metabolism. Omega-3 polyunsaturated fatty acids (n-3 PUFA) have anti-inflammatory activity and suppresses hepatic triacylglycerol accumulation, but many daily diets are deficient in this nutrient. Therefore, the effect of DE exposure in mice fed n-3 PUFA-deficient diet was investigated. Mice were fed control chow or n-3 PUFA-deficient diet for 4 weeks, then exposed to clean air or DE by inhalation for further 4 weeks. Liver histology, plasma parameters, and expression of fatty acid synthesis-related genes were evaluated. N-3 PUFA-deficient diet increased hepatic lipid droplets accumulation and expression of genes promoting fatty acid synthesis: Acaca, Acacb, and Scd1. DE further increased the plasma leptin and the expression of fatty acid synthesis-related genes: Acacb, Fasn, and Scd1. N-3 PUFA-deficient diet and DE exposure potentially enhanced hepatic fatty acid synthesis and subsequently accumulation of lipid droplets. The combination of low-dose DE exposure and intake of n-3 PUFA-deficient diet may be an additional risk factor for the incidence of non-alcoholic fatty liver disease. The present study suggests an important mechanism for preventing toxicity of DE on the liver through the incorporation of n-3 PUFAs in the diet.

Separation of cultured strawberry cells producing anthocyanins in aqueous two-phase system.

A rapid and simple selection method of high-yield cells has been desired to establish highly productive cell lines for useful secondary metabolites. For this purpose, a new attempt was made to partition cultured plant cells in a poly(ethylene glycol)-dextran aqueous two-phase system (ATPS). The applicability of the ATPS in partitioning cultured strawberry cells (designated FAW) was investigated. The result of single-step partitioning in the ATPS supplemented with 0.4 mmol/kg lithium sulfate showed that FAW cells cultivated for 7 d under light-irradiation were separated into two cell populations with significantly different anthocyanin content. Additionally, the analysis technique of microscopic cell images showed that cells accumulating a high level of anthocyanin were partitioned completely into the bottom phase in a partitioning experiment of FAW cells cultivated for 10 d under light-irradiation in the ATPS supplemented with 1.8 mmol/kg potassium phosphate buffer. These results indicated that cell partitioning in ATPS increased the intracellular anthocyanin content and that the cultured strawberry cell population was heterogeneous in terms of cell surface properties. This is the first report of partitioning based on the heterogeneity of the cell surface properties correlated with the intracellular secondary metabolism in cultured plant cells. Our results also suggested that the ATPS was appropriate as a large-scale method for selecting useful cell lines among the cultured plant cells.

Compounds blocking mutant huntingtin toxicity identified using a Huntington's disease neuronal cell model.

Neuronal cell death in HD is believed to be largely a dominant cell-autonomous effect of the mutant huntingtin protein. We previously developed an inducible PC12 cell model which expresses an N-terminal huntingtin fragment with an expanded poly Q repeat (N63-148Q) under the control of the tet-off system. In order to evaluate the ability of compounds to protect against mutant huntingtin toxicity in our model, we measured LDH released by dead cells into the medium. We have now screened the library of 1040 compounds from the NINDS Custom Collection as part of a National Institute of Neurological Disorders and Stroke (NINDS) collaborative project. Each positive compound was tested at 3-8 concentrations. Five compounds significantly attenuated mutant huntingtin (htt)-induced LDH release without affecting the expression level of huntingtin and independent of effect on aggregates. We also tested a broad spectrum caspase inhibitor Z-VAD-fmk and previously proposed candidate compounds. This cell model can provide a method to screen potential therapeutic compounds for treating Huntington's disease.

Positional candidate approach for the gene responsible for benign adult familial myoclonic epilepsy.

Benign adult familial myoclonic epilepsy (BAFME) is an autosomal dominant idiopathic epileptic syndrome characterized by adult-onset tremulous finger movement, myoclonus, epileptic seizures, and nonprogressive course, recognized in Japanese families. We recently assigned the gene locus to chromosome 8q23.3-q24.11 by linkage analysis. In this study, we identified the sequence of human cDNA encoding Kv8.1, a neuronal modulatory alpha-subunit of the voltage-gated potassium channel, mapped to human chromosome 8q22.3-8q24.1. Human Kv8.1 cDNA had a 1,500-nucleotide open reading frame encoding a polypeptide of 500 amino acids, in which six hydrophobic transmembrane segments were well conserved, and its overall amino acids homology as compared with those of rat and golden hamster was 95.0% and 95.0%. Tissue-distribution analysis by reverse transcription-polymerase chain reaction (RT-PCR) showed the presence of the Kv 8.1 transcript exclusively in the brain. The analysis of the genomic organization of the human gene revealed consistency of three exons interrupted by two introns each of 1.2 and 3.5 kb. We analyzed the genomic sequence of the patients with BAFME and found no change in the pathogenesis of the disease.