Apolipoprotein E allele-specific antioxidant activity and effects on cytotoxicity by oxidative insults and beta-amyloid peptides.

The apolipoprotein E (APOE) E4 allele is associated with Alzheimer's disease, cardiovascular disease, and decreased longevity. To probe the mechanism of these associations, cell lines were created which secrete each apoE isoform. ApoE conditioned media, purified apoE, and commercially obtained apoE protected B12 cells from hydrogen peroxide cytotoxicity with E2 > E3 > E4. Physiological levels of apoE protected cells from beta-amyloid peptides, while higher doses of apoE led to increased cytotoxicity. E2 > E3 > E4 possessed antioxidant activity, and apoE bound certain metal ions. The decreased antioxidant activity of E4 could contribute to its association with Alzheimer's disease, cardiovascular disease and decreased longevity.

The effect of Daikenchuto on blood flow of the superior mesenteric artery and portal vein in ELBW: A prospective study.

BACKGROUND: Focal intestinal perforation (FIP) is a devastating complication of premature birth, and extremely low birth weight (ELBW) infants are at highest risk. This study aimed to evaluate the relationship of the superior mesenteric artery (SMA) and portal vein (PV) blood flow velocities to investigate the association between intestinal blood flow and FIP. In addition, the herbal formula Daikenchuto (TJ-100) is expected to improve intestinal blood flow disorders; therefore, we evaluated its effect. METHODS: We conducted a prospective cohort study of 15 ELBW infants from January 2020 to August 2021. Measured variables included birth weight, 5-minute Apgar score, time of oral feeding initiation, ductus arteriosus (PDA) closure (percent), diastolic and systolic blood pressure, SMA and PV blood flow velocity, and FIP onset data. Fifteen infants were divided into three groups: a non-surgery group (Group I; 6), a surgery group with FIP (Group II; 4), and a TJ-100 administration group (Group III; 5). The main outcome parameters included SMA and PV blood flow velocities with TJ-100. RESULTS: SMA and PV blood flow differed significantly for the SMA of Group I and the SMA and PV of Group III (P < 0.01, P = 0.01, and P = 0.04, respectively). There was a correlation between SMA and PV in Group III (P = 0.03). CONCLUSION: TJ-100 may increase SMA and PV blood flow and improve intestinal blood flow in ELBW infants at risk of FIP. Therefore, the effects of TJ-100 should undergo further study.

Electrochemical oxidation of tetracycline antibiotics using a Ti/IrO2 anode for wastewater treatment of animal husbandry.

In animal husbandry, antibiotics are widely used to treat and prevent diseases or to promote growth. The use of antibiotics for domestic animals enables to promote safety of livestock products and enhance productivity. Tetracycline antibiotics (TCs) are one of the primarily used groups of antibiotics for cattle and swine. However, the unintentional spreading of antibiotics from animal waste to the environment may leave out drug residues, promoting resistant strains of bacteria, and will adversely affect the ecosystem and human health. To prevent the spread of veterinary antibiotics in the environment, it is required to treat residual antibiotics in livestock wastewater. In this study, we investigated the electrochemical oxidation of TCs to treat livestock wastewater. The concentrations of TCs in aqueous solutions were reduced from 100 mg/L to less than 0.6 mg/L by 6 h of electrochemical treatment using a Ti/IrO2 anode with Na2SO4 electrolyte. The concentration of oxytetracycline (OTC) in livestock wastewater was also reduced from 100 mg/L to less than 0.7 mg/L by the same treatment. Thus, the electrochemical oxidation using a Ti/IrO2 anode with Na2SO4 electrolyte was found to be effective for degradation of TCs. The results suggest that the electrochemical oxidation method is a promising treatment for TCs in livestock wastewater.

A case of neonate effectively treated with everolimus for giant hepatic hemangioma complicated with congenital duodenal atresia and Kasabach-Merritt syndrome.

BACKGROUND: Disseminated intravascular coagulation (DIC) with Kasabach-Merrit syndrome from a large hepatic hemangioma is life-threatening. We report a case of giant hepatic hemangioma of the newborn with KMS. RESULTS: The patient was born at 37 gestational weeks and 2 days via cesarean section; weight at birth was 2952 g. Congenital duodenal atresia was noted during the fetal period. DIC developed after delivery and a giant liver hemangioma was diagnosed via abdominal CT. The cause of DIC was Kasabach-Merritt syndrome owing to a giant hepatic hemangioma. First, combination therapy of 2 mg/kg/day of prednisolone and 0.2 mg/kg/day of propranolol was initiated form enterostomy. However, the size of the hepatic hemangioma did not alter, as observed via image evaluation. Therefore, 0.3 mg/kg/day of everolimus was administered frorm enterostomy. Subsequently, the size of the hepatic hemangioma was assessed via image evaluation. Although it did not alter, blood flow to the hepatic hemangioma decreased and thrombocytopenia was also suppressed. We performed hepatic lateral segmentectomy, radical operation for duodenal atresia. The pathological diagnosis of the removed tumor was infantile hemangioma. CONCLUSION: We report everolimus may be useful when PSL and propranolol are ineffective.

Intercalation of alkylamines into an organic polymer crystal

Organic solid-state synthesis allows formation of products that are difficult or impossible to produce by conventional methods. This feature, and the high degree of reaction selectivity that can be achieved, is a direct result of the control over the relative orientation of the reactants afforded by the solid state. But as the successful development of 'topochemical reactions' requires the careful design of suitable reactant crystals, the range of both reactions and products amenable to this approach has been limited. However, recent advances in organic crystal engineering, particularly the rational design of complex solid architectures through supramolecular preorganization, have renewed interest in topochemical reactions. Previously, we have orientated muconate monomers--diene moieties with a carboxylate group on each end--using long-chain n-alkylammonium ions, such that the topochemical photopolymerization of the solid-state reactants produces layered crystals of stereoregular and high-molecular-mass polymers. Here we show that these polymer crystals are capable of repeated, reversible intercalation by conversion to the analogous poly(carboxylic acid), followed by transformation into a number of poly(alkylammonium muconate)s upon addition of the appropriate amine. Introduction of functional groups into these crystals may allow the design of organic solids for applications such as molecular recognition, separation and catalysis, thereby extending the range and practical utility of current intercalation compounds.

Construction of a model cell line for the assay of MDR1 (multi drug resistance gene-1) substrates/inhibitors using HeLa cells.

Cancer cells often become resistant to chemotherapy, and induction of the ABC transporter Multi-drug Resistance gene-1 (MDR1) is a major cause. We established a tool for high-throughput screening of substrates and inhibitors of MDR1, using transformed HeLa cells that over-express MDR1. The cDNA for human MDR1 was subcloned into the eukaryotic expression vector pBK-CMV to produce an MDR1 expression vector, pBK-CMV/MDR1. HeLa cells were transfected with pBK-CMV/MDR1 or the empty vector pBK-CMV. Transfection of the vector sequence for MDR1 and its expression were evaluated by genomic PCR and western blotting, respectively. The efficiency of the MDR1 transporter for pumping a substrate out of the transformed cells was evaluated using rhodamine123 (R-123), a mitochondrial dye that is also an MDR1 substrate. After treatment of the MDR1-expressing HeLa cells with MDR1 substrate vinblastin or inhibitors cyclosporin A and verapamil, the amount of R-123 retained in the cells was increased to 2 to 2.3 times the level in untreated MDR1-expressing HeLa cells. The transfection of empty pBK-CMV had no effect on the R-123 retention in HeLa cells, regardless of drug treatment. In conclusion, we have established a model human carcinoma cell line that expresses functional MDR1 and can be used to screen for substrates and inhibitors of MDR1.

Corticotropin-releasing factor plays a permissive role in cerebellar long-term depression.

This study of rat cerebellar slices yielded two lines of evidence indicating that the corticotropin-releasing factor (CRF) found in climbing fibers (CFs) is critical for the induction of long-term depression (LTD) at the parallel fiber (PF) synapses of Purkinje cells (PCs) by their conjunctive activation with either stimulation of CFs or depolarization of PCs. First, LTD induction was effectively blocked by specific CRF receptor antagonists, alpha-helical CRF-(9-41) (alpha-h CRF) and astressin; and second, LTD was no longer observed in CF-deprived cerebella but was restored by CRF replenishment. The data obtained in this study suggest that these effects are mediated by protein kinase C (PKC) and not by Ca2+ signaling or cyclic GMP (cGMP) production.

Local calcium release in dendritic spines required for long-term synaptic depression.

We have used rats and mice with mutations in myosin-Va to evaluate the range and function of IP3-mediated Ca2+ signaling in dendritic spines. In these mutants, the endoplasmic reticulum and its attendant IP3 receptors do not enter the postsynaptic spines of parallel fiber synapses on cerebellar Purkinje cells. Long-term synaptic depression (LTD) is absent at the parallel fiber synapses of the mutants, even though the structure and function of these synapses otherwise appear normal. This loss of LTD is associated with selective changes in IP3-mediated Ca2+ signaling in spines and can be rescued by photolysis of a caged Ca2+ compound. Our results reveal that IP3 must release Ca2+ locally in the dendritic spines to produce LTD and indicate that one function of dendritic spines is to target IP3-mediated Ca2+ release to the proper subcellular domain.

Clock gene dysfunction in patients with obstructive sleep apnoea syndrome.

Clock genes regulate mammalian circadian rhythms, and dysfunction of clock genes can contribute to various disorders. To investigate whether obstructive sleep apnoea syndrome (OSAS) influences clock gene function, the present authors examined Period1 (Per1) mRNA expression in vitro and in vivo. In eight healthy subjects and eight OSAS patients, plasma noradrenaline, serum interleukin (IL)-6, high-sensitivity C-reactive protein (hsCRP) and Per1 mRNA expression in peripheral whole blood were measured. Expression of Per1 mRNA in cultured cells was examined under IL-6 or noradrenaline stimulation in vitro. After noradrenaline was administered to mice in vivo, Per1 mRNA expression in the brain was examined. The concentrations of serum IL-6, hsCRP and plasma noradrenaline were elevated in OSAS patients, but improved by continuous positive airway pressure (CPAP) therapy. Per1 mRNA expression in the peripheral blood significantly decreased at 02:00 h by CPAP in OSAS patients. Stimulation with IL-6 did not directly induce Per1 mRNA in vitro. Administration of noradrenaline induced Per1 mRNA in the cerebral cortex of mice in vivo. The current study revealed that obstructive sleep apnoea syndrome caused clock gene dysfunction, and continuous positive airway pressure helped to improve it. Sympathetic activation and elevation of the plasma noradrenaline concentration in obstructive sleep apnoea syndrome may be one of the factors involved in disorders of Period1 mRNA expression.

Actinobacillus actinomycetemcomitans lipopolysaccharide stimulates collagen phagocytosis by human gingival fibroblasts.

INTRODUCTION: Collagen phagocytosis by fibroblasts is involved in the intracellular pathway related to collagen breakdown in soft connective tissues. The possible role of lipopolysaccharide (LPS) in regulating this fibroblast function has not been elucidated so we investigated the effect of LPS from Actinobacillus actinomycetemcomitans, a periodontopathic bacterium, on collagen phagocytic activity in human gingival fibroblasts and associated regulatory mechanisms. METHODS: LPS pretreatment stimulated binding of collagen-coated beads to cells and, subsequently, their internalization. RESULTS: The LPS-activated collagen phagocytic process was enhanced in the presence of the soluble form of CD14 (sCD14) or LPS-binding protein (LBP), while the LPS/LBP treatment activated Akt and induced actin reorganization. Furthermore, these LPS/LBP-induced effects were partially suppressed by adding phosphatidyl-inositol-3 kinase (PI3K) inhibitors. CONCLUSION: These results suggest that A. actinomycetemcomitans LPS disturbs the homeostasis of collagen metabolism within gingival tissue by facilitating collagen phagocytosis by gingival fibroblasts, and serum sCD14 and LBP positively regulate the action of LPS. In addition, the PI3K/Akt signaling is thought to partially mediate the LPS/LBP-stimulated collagen phagocytic pathway, which may be dependent on actin cytoskeletal rearrangement.

On the search for markers of tick resistance in bovines.

Genetic differences in susceptibility to ticks (Rhipicephalus (Boophilus) microplus) are considerable in bovines. Here, mapping, association and gene expression approaches were employed to further advance our understanding of the molecular basis of tick resistance. A B. taurus x B. indicus F2 population was developed by Embrapa and 382 individuals were measured for parasitic load. Scanning of all chromosomes is in progress. Quantitative trait loci (QTL) for tick load were mapped to chromosomes 4, 5, 7, 10, 14, 18 and 23 out of the 20 chromosomes scanned and were dependent on the season in which the phenotype was scored. In the candidate gene approach, females from the genetic groups Nelore (NE--184), Canchim x Nelore (CN--153), Aberdeen Angus x Nelore (AN--123) and Simmental x Nelore (SN--120) were evaluated under natural infestation. Microsatellite markers close to the genes for interleukin 2 (IL2), interleukin 4 (IL4) and interferon gamma (IFNG) were analysed. Tick counts were associated with the marker for interleukin 4 (P < 0.05) in three genetic groups. Differences in cytokine mRNA levels of naive versus infested Nelore calves as well as between resistant versus susceptible cows from NE, CN and AN genetic groups were also investigated. Comparison of cytokines from infested and naïve animals showed downregulation of IL2. When resistant cows were compared to susceptible animals, IL8 was downregulated. These results reinforce the multiloci nature of tick resistance and the need to consider QTL and environment interactions.

Effects of assisted reproductive technologies in neonates with indications for surgery (2007-2016).

PURPOSE: Recently, the number of births using assisted reproductive technologies (ART) has increased. An associated increase in the incidence of congenital malformations in babies conceived using this technology has also been reported. Therefore, we aimed to investigate the rate of malformations in babies with neonatal surgical diseases, who were conceived using ART. MATERIALS AND METHODS: Between January 2007 and December 2016, 1737 patients were admitted to our hospital. We analyzed the incidence of congenital cardiac diseases, genetic anomalies, and congenital anomalies of the kidney and urinary tract (CAKUT) in neonates conceived by ART. The χ2 test and logistic regression analysis were used to assess the odds ratios (ORs) for congenital malformations. A P-value < 0.05 indicated statistical significance. RESULTS: The OR for CAKUT was 16.94 for the first-birth neonates conceived using ART, [P < 0.05, AUC (area under the curve) = 0.86]. However, for non-surgery neonates, the OR for CAKUT was 5.99 (P = 0.15, AUC = 0.87), compared to 32.27 (P < 0.05, AUC = 0.93) for parallel conditions in surgery-neonates. CONCLUSION: Neonates conceived using ART are prone to develop CAKUT, which will need surgical treatment. Therefore, more management is necessary for associated malformations in these babies, particularly in cases with CAKUT.

Signal Change of Acute Cortical and Juxtacortical Microinfarction on Follow-Up MRI.

BACKGROUND AND PURPOSE: Although the clinical importance of cortical microinfarcts has become well-recognized recently, the evolution of cortical microinfarcts on MR imaging is not fully understood. The aim of this study was to examine the temporal changes in acute cortical microinfarcts using susceptibility-weighted imaging and conventional MR imaging. MATERIALS AND METHODS: Patients with acute infarcts located in the cortical and/or juxtacortical region measuring ≤10 mm in axial diameter based on diffusion-weighted imaging who had a follow-up 3T MR imaging were retrospectively included in the study. All lesions did not show hypointensity on initial T2*WI. For cortical and/or juxtacortical microinfarcts detected on initial DWI, 2 neuroradiologists evaluated the follow-up MR imaging (T2WI, FLAIR, T2*WI, and SWI) and assessed lesion signal intensities and locations (cortical microinfarcts or microinfarcts with juxtacortical white matter involvement). RESULTS: On initial DWI, 2 radiologists observed 180 cortical and/or juxtacortical microinfarcts in 35 MR imaging examinations in 25 patients; on follow-up, the neuroradiologists identified 29 cortical microinfarcts (16%) on T2WI, 9 (5%) on FLAIR, 4 (2%) on T2*, and 97 (54%) on SWI. All cortical microinfarcts detected with any follow-up MR imaging showed hyperintensity on T2WI/FLAIR and/or hypointensity on T2*WI and SWI. CONCLUSIONS: SWI revealed conversion (paramagnetic susceptibility changes) of acute cortical microinfarcts, suggesting that a substantial number of cortical microinfarcts may contain hemorrhagic components.

Pre-Multidrug-Resistant Mycobacterium tuberculosis Infection Causing Fatal Enteric Disease in a Dog from a Family with History of Human Tuberculosis.

This report describes a fatal case of a pet dog with major enteric signs owned by a family that has experienced cases of pulmonary tuberculosis (TB) in the household. Clinical and epidemiological aspects, imaging data, microbiological, haematological and histopathological examinations were assessed to diagnosis of disease. gyrB-RFLP, spoligotyping and MIRU-VNTR allowed molecular detection of M. tuberculosis strain from S family. The resazurin microtiter assay indicated that all isolates were resistant to isoniazid, ethambutol, ciprofloxacin, ofloxacin, streptomycin and amikacin. The public health concerns related to canine tuberculosis and risk of the dissemination by pets of M. tuberculosis pre-multidrug-resistant (PMD) to isoniazid, ethambutol and other first-line drugs used in human therapy of TB are discussed. We believe this to be the first report of PMD M. tuberculosis infection in a dog presenting mainly enteric manifestation, confirmed as S lineage by molecular methods, owned by a family in which TB has spread in the household for generations.

In vitro potentiation of human natural killer cell activity by a streptococcal preparation, OK-432: interferon and interleukin-2 participation in the stimulation with OK-432.

Inasmuch as human natural killer (NK) cell activity was markedly augmented by a streptococcal immunopotentiator, OK-432, both in vivo and in vitro, the mechanism in which OK-432 augmented human NK cell activity was analyzed. Culture supernatants of nonadherent lymphocytes stimulated with OK-432 significantly augmented NK cell activity. Significant activity of both interferon (IFN) (both alpha- and gamma-types) and interleukin-2 (IL-2) was detected in the culture supernatants from nonadherent lymphocytes. Concomitant treatment of supernatants with anti-IFN-alpha antiserum and pH-2 glycine-HCI buffer or the absorption of supernatants with an IL-2-dependent cell line completely abrogated the NK-augmenting activity, whereas the treatment with either one of these resulted in only partial elimination of the activity. These results indicate that OK-432 stimulates human nonadherent lymphocytes to produce IFN and IL-2 and that both factors are primarily responsible for the NK augmentation by OK-432.

Inner segment ellipsoid band length is a prognostic factor in retinitis pigmentosa associated with EYS mutations: 5-year observation of retinal structure.

PurposeTo evaluate whether the length of the inner segment ellipsoid (ISe) band can be used as a prognostic factor for disease course in retinitis pigmentosa (RP) patients with EYS mutations by observation over a period of 5 years.MethodsTwelve RP patients with EYS mutations were studied. The horizontal and vertical ISe length of the right eye was manually measured at five time points annually, using spectral domain optical coherence tomography. A regression line through the five points from baseline to the final measurement was drawn and the ratio of the length (%) at each point to the baseline length was calculated; the slope was defined as the rate of ISe shortening (%/year). The correlation between the rate of ISe shortening and age, visual acuity, and mean deviation (MD) value were evaluated. The intraclass correlation coefficient (ICC) for the measurements was calculated.ResultsThe mean rate of ISe shortening was -4.65±2.89% per year and the decline was statistically significant. The rate of shortening was significantly negatively correlated with the baseline length (P=0.046, r=0.58), but not with the baseline age, visual acuity, and MD value. The ICC (2, 1) was 0.999.ConclusionsISe of all RP patients with EYS mutations shortened during the 5 years of annual observation. The measurement of the length of ISe is a simple and convenient method with high repeatability, and the length is a sensitive prognostic factor for the rate of ISe shortening in RP patients with EYS mutations.

Extensive use of vasodilator agents and functional echocardiography to monitor extremely-low-birth-weight infants in Japan.

National surveys were conducted in Japan to assess the current practices for circulatory management of extremely-low-birth-weight infants (ELBWIs) in acute phases. Approximately 80 and 100 institutions were surveyed in 2006 and 2011, respectively. Echocardiography was identified as an important diagnostic tool at 95% of the surveyed institutions. Furthermore, 74% of the institutions survey in 2011 used vasodilator agents. In 2011, the mean velocity of circumferential fiber shortening (mVcfc) and left ventricular end-systolic wall stress (ESWS) were used by 60% of the surveyed institutions to evaluate the relationship between afterload of the left ventricle and left ventricular contractility. Overall, the data collected from these national surveys clarified the current practices for circulatory management of ELBWIs in Japan, particularly the use of echocardiography and cardiovascular agents, including catecholamines and vasodilators.

Distinct roles of Galpha(q) and Galpha11 for Purkinje cell signaling and motor behavior.

G-protein-coupled metabotropic glutamate group I receptors (mGluR1s) mediate synaptic transmission and plasticity in Purkinje cells and, therefore, critically determine cerebellar motor control and learning. Purkinje cells express two members of the G-protein G(q) family, namely G(q) and G11. Although in vitro coexpression of mGluR1 with either Galpha11 or Galpha(q) produces equally well functioning signaling cascades, Galpha(q)- and Galpha11-deficient mice exhibit distinct alterations in motor coordination. By using whole-cell recordings and Ca2+ imaging in Purkinje cells, we show that Galpha(q) is required for mGluR-dependent synaptic transmission and for long-term depression (LTD). Galpha11 has no detectable contribution for synaptic transmission but also contributes to LTD. Quantitative single-cell RT-PCR analyses in Purkinje cells demonstrate a more than 10-fold stronger expression of Galpha(q) versus Galpha11. Our findings suggest an expression level-dependent action of Galpha(q) and Galpha11 for Purkinje cell signaling and assign specific roles of these two G(q) isoforms for motor coordination.

Head-to-head linkage of carp alpha- and beta-globin genes.

The alpha- and beta-globin gene variants are believed to have diverged from a single ancestral globin gene, and the divergence was primed by the duplication of the ancestral globin gene. To understand the process of gene duplication, we investigated the alpha- and beta-globin gene arrangement of a bony fish (carp). From a Southern analysis of seven previously prepared lambda phage clones (lambdaCG1-7) using radio-labelled alpha- or beta-globin gene probes, it was found that the clones included both the alpha- and beta-globin genes, and that they were located within a distance of 1 kb. Additionally, the linkage of two alpha-globin genes and two beta-globin genes in the clone lambdaCG1, 5 and 7 (e.g., alpha-beta-alpha-beta in lambdaCG5) revealed an arrangement that is different from the arrangement in higher vertebrates in which the alpha-globin and beta-globin genes generally occur at different loci. The distances between the detached alpha- to beta-globin genes were approximately 5 to 10 kb. DNA sequencing of the adjacently linked alpha- and beta-globin genes in lambdaCG3 showed that they were arranged in a head-to-head orientation. PCR amplification using primers for the internal region between the carp alpha- and beta-globin genes gave approximately 0.9-kb products from each of the clones lambdaCG1, 3, 4, 5, 6, and 7, and from the chromosomal DNA of German mirror carp, Saku carp, Suwa carp, and Yamato carp. This demonstrates the alternative arrangement of carp alpha- and beta-genes in the globin gene locus (i.e., 3'alpha5'-5'beta3'-3'alpha5'-5'beta3' in lambdaCG5), and the widespread distribution of head-to-head-linked alpha- and beta-globin genes in carp. Based on the above results, we hypothesize that the duplication of the ancestral globin gene (prior to the divergence of the a and beta forms) occurred in a head-to-head direction.

Identification of cAMP analogue inducible genes in RAW264 macrophages.

RNA was isolated from RAW264 cells treated with or without 8-Br-cAMP and the differential display and subtractive hybridization methods were performed. One hundred and twenty-five differentially displayed bands were identified. Upon Northern blot analysis, only three of these bands were confirmed as cAMP inducible mRNAs, named cI-1, cI-2, and cI-3 (for cAMP inducible genes 1-3). The cI-3 probe was identical to a previously known gene, gly96. Using the novel cI-1 and cI-2 partial cDNAs as probes, a mouse macrophage cDNA library was screened and the two full length genes were cloned, sequenced, and characterized as encoding large hydrophobic proteins. One hundred and fifteen partial cDNA clones from a subtractive hybridization library were also screened by Northern blot and 64 were found to be cAMP inducible. Of these, 45 represented 31 known unique genes in the GenBank nr database (cI-4-34), and 19 clones representing 15 unique sequences were not in the nr database (cI-35-49). One of the previously known genes was ABC1, the Tangier disease gene, which was identified from four independent partial cDNAs. ABC1 was upregulated in RAW cells by cAMP, concurrent with the cAMP induction of lipid efflux to apolipoprotein A1.