A Hybrid Auricular Framework of Autologous Rib Cartilage and a Porous Polyethylene Implant for Reconstruction of Congenital Microtia: A Modification of Nagata's Technique.

Background: An implant (porous polyethylene) is an alternative to rib cartilage for microtia reconstruction but carries a risk of extrusion. Objective: To evaluate the outcome of a hybrid framework of implant with rib cartilage for microtia reconstruction. Methods: Patients who underwent Nagata's technique for microtia reconstruction were reviewed for complications and aesthetic score. In stage 1, a rib cartilage framework or a hybrid framework of implant with rib cartilage was used. In stage 2, the framework was elevated and supported by an implant for projection. Postoperative outcomes were reported for both groups. Results: Forty-four ears of 40 patients underwent surgery. Eleven ears received a rib auricular framework and 33 ears a hybrid auricular framework. The mean postoperative follow-up for the rib and hybrid groups was 76.3 and 43.1 months, respectively. No supporting postauricular implant extruded, whereas stainless-steel wires extruded in seven ears (15.9%). Five (15.2%) hybrid frameworks were removed due to infection or extrusion. Mean operating time was 2 h shorter in the hybrid group. Aesthetic outcomes were similar for both groups. Conclusion: A hybrid framework of rib and implant that requires less harvested cartilage is feasible for microtia reconstruction, but caution should be used due to its higher explantation rate.

Regulation of proton partitioning in kinase-activating acute myeloid leukemia and its therapeutic implication.

Gain-of-function kinase mutations are common in AML and usually portend an inferior prognosis. We reported a novel mechanism whereby kinase mutants induced intracellular alkalization characteristic in oncogenesis. Thirteen kinases were found to activate sodium/hydrogen exchanger (NHE1) in normal hematopoietic progenitors, of which FLT3-ITD, KRASG12D, and BTK phosphorylated NHE1 maintained alkaline intracellular pH (pHi) and supported survival of AML cells. Primary AML samples with kinase mutations also showed increased NHE1 phosphorylation and evidence of NHE1 addiction. Amiloride enhanced anti-leukemic effects and intracellular distribution of kinase inhibitors and chemotherapy. Co-inhibition of NHE1 and kinase synergistically acidified pHi in leukemia and inhibited its growth in vivo. Plasma from patients taking amiloride for diuresis reduced pHi of leukemia and enhanced cytotoxic effects of kinase inhibitors and chemotherapy in vitro. NHE1-mediated intracellular alkalization played a key pathogenetic role in transmitting the proliferative signal from mutated-kinase and could be exploited for therapeutic intervention in AML.

Function of Arl4aa in the Initiation of Hematopoiesis in Zebrafish by Maintaining Golgi Complex Integrity in Hemogenic Endothelium.

ADP-ribosylation factor-like 4aa (Arl4aa) is a member of the ADP-ribosylation factor family. It is expressed in hematopoietic tissue during embryonic development, but its function was unknown. Zebrafish arl4aa is preferentially expressed in the ventral wall of the dorsal aorta (VDA) at 24 and 36 hpf and in caudal hematopoietic tissue at 48 hpf. Morpholino knockdown and transcription activator-like effector nuclease (TALEN) knockout of arl4aa significantly reduced expression of genes associated with definitive hematopoietic stem cells (HSCs). Golgi complex integrity in VDA was disrupted as shown by transmission electron microscopy and immunostaining of Golgi membrane Giantin. Mechanistically, arl4aa knockdown reduced Notch signaling in the VDA and its target gene expression. Protein expression of NICD was also reduced. Effects of arl4aa knockdown on definitive hematopoiesis could be restored by NICD expression. This study identified arl4aa as a factor regulating initiation of definitive HSCs by maintaining the integrity of Golgi complex and, secondarily, maturation of the Notch receptor.

Sorafenib and omacetaxine mepesuccinate as a safe and effective treatment for acute myeloid leukemia carrying internal tandem duplication of Fms-like tyrosine kinase 3.

BACKGROUND: Omacetaxine mepesuccinate (OME) has antileukemic effects against acute myeloid leukemia (AML) carrying an internal tandem duplication of Fms-like tyrosine kinase 3 (FLT3-ITD). A phase 2 clinical trial was conducted to evaluate a combination treatment of sorafenib and omacetaxine mepesuccinate (SOME). METHODS: Relapsed or refractory (R/R) or newly diagnosed patients were treated with sorafenib (200-400 mg twice daily) and OME (2 mg daily) for 7 (first course) or 5 days (second course onward) every 21 days until disease progression or allogeneic hematopoietic stem cell transplantation (HSCT). The primary endpoint was composite complete remission, which was defined as complete remission (CR) plus complete remission with incomplete hematologic recovery (CRi). Secondary endpoints were leukemia-free survival (LFS) and overall survival (OS). RESULTS: Thirty-nine R/R patients and 5 newly diagnosed patients were recruited. Among the R/R patients, 28 achieved CR or CRi. Two patients showed partial remission, and 9 patients did not respond. Among the 5 newly diagnosed patients, 4 achieved CR, and 1 achieved CRi. The median LFS and OS were 5.6 and 10.9 months, respectively. Prior Fms-like tyrosine kinase 3 (FLT3) inhibitor exposure (P = .007), 2 or more inductions (P = .001), and coexisting IDH2 (P = .008) and RUNX1 mutations (P = .003) were associated with lower CR/CRi rates. HSCT consolidation and deep molecular responses (defined as an FLT3-ITD variant allelic frequency [VAF] ≤ 0.1% or a nucleophosmin 1 [NPM1] mutant VAF ≤ 0.01%) were associated with better OS and LFS. Prior FLT3 inhibitor exposure and 2 or more inductions were associated with inferior LFS. CONCLUSIONS: SOME was safe and effective for R/R and newly diagnosed FLT3-ITD AML.

Distinct mutation spectrum, clinical outcome and therapeutic responses of typical complex/monosomy karyotype acute myeloid leukemia carrying TP53 mutations.

The present study aimed to define a subtype of complex/monosomal karyotype (CK/MK) acute myeloid leukemia (AML) by its distinct clinical features, p53 signaling and responses to p53 targeting agents. Ninety-eight young adults (range: 21-60 years; median: 49 years) with CK/MK AML were studied. They received standard induction, consolidation and allogeneic hematopoietic stem cell transplantation from siblings or matched unrelated donors if available. Chromosomal abnormalities most commonly affected chromosome 5 (30%), 7 (22%) and 17 (21%). Next generation sequencing of a 54-myeloid gene panel were available in 76 patients. Tumor protein 53 (TP53) mutations were most common (49%) and associated with the presence of -5/5q- (P < .001) and -17/17p- (P < .001), but not -7/7q- (P = .370). This "typical" CK/MK AML subtype was associated with significantly lower presenting white cell counts, higher number of karyotypic abnormalities, and inferior leukemia-free and overall survivals, compared with CK/MK AML without the typical features. Blood or bone marrow samples from typical CK/MK AML patients showed defective p53 signaling upon induction by etoposide. In vitro drug sensitivity analysis showed that they were sensitive to APR-246 that targeted mutant p53, but resistant to MDM2 antagonist MI-77301. Novel therapeutic strategies targeting TP53 mutations in CK/MK AML should be developed and tested in clinical trials.

Targeted sequencing of ABCA7 identifies splicing, stop-gain and intronic risk variants for Alzheimer disease.

Several variants in the gene ABCA7 have been identified as potential causal variants for late-onset Alzheimer's disease (LOAD). In order to replicate these findings, and search for novel causal variants, we performed targeted sequencing of this gene in cohorts of non-Hispanic White (NHW) and African-American (AA) LOAD cases and controls. We sequenced the gene ABCA7 in 291 NHW LOAD cases and 103 controls. Variants were prioritized for rare, damaging variants and previously reported variants associated with LOAD, and were follow-up genotyped in 4076 NHW and 1157 AA cases and controls. We confirm three previously associated ABCA7 risk variants and extend two of these associations to other populations, an intronic variant in NHW (P=3.0×10-3) (originally reported in a Belgian population), and a splice variant originally associated in the Icelandic population, which was significantly associated in the NHW cohort (P=1.2×10-6) and nominally associated in the AA cohort (P=0.017). We also identify a 3'-UTR splice variant that segregates in four siblings of one family and is nominally associated with LOAD (P=0.040). Multiple variants in ABCA7 contribute to LOAD risk.

Homoharringtonine (omacetaxine mepesuccinate) as an adjunct for FLT3-ITD acute myeloid leukemia.

An in vitro drug-screening platform on patient samples was developed and validated to design personalized treatment for relapsed/refractory acute myeloid leukemia (AML). Unbiased clustering and correlation showed that homoharringtonine (HHT), also known as omacetaxine mepesuccinate, exhibited preferential antileukemia effect against AML carrying internal tandem duplication of fms-like tyrosine kinase 3 (FLT3-ITD). It worked synergistically with FLT3 inhibitors to suppress leukemia growth in vitro and in xenograft mouse models. Mechanistically, the effect was mediated by protein synthesis inhibition and reduction of short-lived proteins, including total and phosphorylated forms of FLT3 and its downstream signaling proteins. A phase 2 clinical trial of sorafenib and HHT combination treatment in FLT3-ITD AML patients resulted in complete remission (true or with insufficient hematological recovery) in 20 of 24 patients (83.3%), reduction of ITD allelic burden, and median leukemia-free and overall survivals of 12 and 33 weeks. The regimen has successfully bridged five patients to allogeneic hematopoietic stem cell transplantation and was well tolerated in patients unfit for conventional chemotherapy, including elderly and heavily pretreated patients. This study validated the principle and clinical relevance of in vitro drug testing and identified an improved treatment for FLT3-ITD AML. The results provided the foundation for phase 2/3 clinical trials to ascertain the clinical efficacy of FLT3 inhibitors and HHT in combination.

A novel Alzheimer disease locus located near the gene encoding tau protein.

APOE ɛ4, the most significant genetic risk factor for Alzheimer disease (AD), may mask effects of other loci. We re-analyzed genome-wide association study (GWAS) data from the International Genomics of Alzheimer's Project (IGAP) Consortium in APOE ɛ4+ (10 352 cases and 9207 controls) and APOE ɛ4- (7184 cases and 26 968 controls) subgroups as well as in the total sample testing for interaction between a single-nucleotide polymorphism (SNP) and APOE ɛ4 status. Suggestive associations (P<1 × 10(-4)) in stage 1 were evaluated in an independent sample (stage 2) containing 4203 subjects (APOE ɛ4+: 1250 cases and 536 controls; APOE ɛ4-: 718 cases and 1699 controls). Among APOE ɛ4- subjects, novel genome-wide significant (GWS) association was observed with 17 SNPs (all between KANSL1 and LRRC37A on chromosome 17 near MAPT) in a meta-analysis of the stage 1 and stage 2 data sets (best SNP, rs2732703, P=5·8 × 10(-9)). Conditional analysis revealed that rs2732703 accounted for association signals in the entire 100-kilobase region that includes MAPT. Except for previously identified AD loci showing stronger association in APOE ɛ4+ subjects (CR1 and CLU) or APOE ɛ4- subjects (MS4A6A/MS4A4A/MS4A6E), no other SNPs were significantly associated with AD in a specific APOE genotype subgroup. In addition, the finding in the stage 1 sample that AD risk is significantly influenced by the interaction of APOE with rs1595014 in TMEM106B (P=1·6 × 10(-7)) is noteworthy, because TMEM106B variants have previously been associated with risk of frontotemporal dementia. Expression quantitative trait locus analysis revealed that rs113986870, one of the GWS SNPs near rs2732703, is significantly associated with four KANSL1 probes that target transcription of the first translated exon and an untranslated exon in hippocampus (P ⩽ 1.3 × 10(-8)), frontal cortex (P ⩽ 1.3 × 10(-9)) and temporal cortex (P⩽1.2 × 10(-11)). Rs113986870 is also strongly associated with a MAPT probe that targets transcription of alternatively spliced exon 3 in frontal cortex (P=9.2 × 10(-6)) and temporal cortex (P=2.6 × 10(-6)). Our APOE-stratified GWAS is the first to show GWS association for AD with SNPs in the chromosome 17q21.31 region. Replication of this finding in independent samples is needed to verify that SNPs in this region have significantly stronger effects on AD risk in persons lacking APOE ɛ4 compared with persons carrying this allele, and if this is found to hold, further examination of this region and studies aimed at deciphering the mechanism(s) are warranted.

Modeling the inactivation of ascaris eggs as a function of ammonia concentration and temperature.

Ammonia sanitization is a promising technology for sanitizing human excreta intended for use as a fertilizer in agriculture. Ascaris eggs are the most persistent pathogens regarding ammonia inactivation and are commonly present in fecal sludge in low- and middle-income countries. In this study, a model for predicting ammonia inactivation of ascaris eggs was developed. Data from four previous studies were compiled and analyzed statistically, and a mathematical model for the treatment time required for inactivation was created. The inactivation rate increased with NH3 activity to the power of 0.7. The required treatment time was found to decrease 10-fold for each 16 °C temperature increase. Dry matter (DM) content and pH had no direct effect on inactivation, but had an indirect effect due to their impact on NH3 activity, which was estimated using the Pitzer approach. An additional model giving an approximation of Pitzer NH3 activity but based on the Emerson approach, DM content and total ammonia (NHTot) was also developed. The treatment time required for different log10 reductions of ascaris egg viability can thus easily be estimated by the model as a function of NH3 activity and temperature. The impact on treatment time by different treatment options can then be theoretically evaluated, promoting improvements of the treatment e.g. by adding urea or alkaline agents, or increasing the temperature by solar heating.

Investigation of quaternary ammonium silane-coated sand filter for the removal of bacteria and viruses from drinking water.

AIMS: To develop an anti-microbial filter media using an attached quaternary ammonium compound (QAC) and evaluate its performance under conditions relevant to household drinking water treatment in developing countries. METHODS AND RESULTS: Silica sand was coated with dimethyloctadecyl [3-(trimethoxysilyl) propyl] ammonium chloride via covalent silane chemistry. Filter columns packed with coated media were challenged with micro-organisms under different water quality conditions. The anti-bacterial properties were investigated by visualizing Escherichia coli (E. coli) attachment to coated media under fluorescence microscopy combined with a live/dead stain. A 9-cm columns with a filtration velocity of 18 m h(-1) achieved log(10) removals of 1·7 for E. coli, 1·8 for MS2 coliphage, 1·9 for Poliovirus type 3 and 0·36 for Adenovirus type 2, compared to 0·1-0·3 log(10) removals of E. coli and MS2 by uncoated sand. Removal scaled linearly with column length and decreased with increasing ionic strength, flow velocity, filtration time and humic acid presence. Escherichia coli attached to QAC-coated sand were observed to be membrane-permeable, providing evidence of inactivation. CONCLUSIONS: Filtration with QAC-coated sand provided higher removal of bacteria and viruses than filtration with uncoated sand. However, major limitations included rapid fouling by micro-organisms and natural organic matter and low removal of viruses PRD1 and Adenovirus 2. SIGNIFICANCE AND IMPACT OF THE STUDY: QAC-coated media may be promising for household water treatment. However, more research is needed on long-term performance, options to reduce fouling and inactivation mechanisms.

An innovative sustainability assessment for urban wastewater infrastructure and its application in Chengdu, China.

Sustainability assessments are an increasingly common tool for measuring progress towards sustainable development. Despite their popularity, sustainability assessments and the indicators that compose them are said to have had little impact on the policy arena. In this paper we discuss four attributes that we contend will improve the use of sustainability assessments to guide decision making: non-compartmentalization, site specificity, built-in guidance for target setting, and ability to measure active sustainability. We present a novel assessment tool for wastewater treatment infrastructure that illustrates these attributes. The assessment is composed of two-dimensional indicators we call "burden to capacity" ratios, that reveal and quantify the local value of resources embodied in wastewater and treatment byproducts, and the tradeoffs between designing systems for disposal versus reuse. We apply the sustainability assessment framework to an existing treatment plant in Chengdu, China and discuss the results.

Recognition and initial management of cardiac emergencies in children.

One may have only minutes to change the trajectory of a child who is deteriorating from either congenital or acquired cardiac disease. However, these children may present with rather cryptic patterns of symptoms (e.g. failure to thrive, lethargy, colic, neonatal shock, respiratory distress, wheezing and syncope with exercise). Thus, it is essential that any health care practitioner who cares for children be familiar with key clinical presentations that require consideration of underlying cardiac disease and time sensitive diagnoses that require rapid recognition and therapy in order to optimize the chances of saving the child's life. The objectives of this manuscript are: 1) to review the initial identification and management of cardiac emergencies in children; and 2) to present a brief summary of key cardiac diagnoses that may need to be considered when caring for children in an acute care setting.

Nutritional management of PKU with glycomacropeptide from cheese whey.

Individuals with phenylketonuria (PKU) must follow a lifelong low-phenylalanine (Phe) diet to prevent neurological impairment. Compliance with the low-Phe diet is often poor owing to restriction in natural foods and the requirement for consumption of a Phe-free amino acid formula or medical food. Glycomacropeptide (GMP), a natural protein produced during cheese-making, is uniquely suited to a low-Phe diet because when isolated from cheese whey it contains minimal Phe (2.5-5 mg Phe/g protein). This paper reviews progress in evaluating the safety, acceptability and efficacy of GMP in the nutritional management of PKU. A variety of foods and beverages can be made with GMP to improve the taste, variety and convenience of the PKU diet. Sensory studies in individuals with PKU demonstrate that GMP foods are acceptable alternatives to amino acid medical foods. Studies in the PKU mouse model demonstrate that GMP supplemented with limiting indispensable amino acids provides a nutritionally adequate source of protein and improves the metabolic phenotype by reducing concentrations of Phe in plasma and brain. A case report in an adult with classical PKU who followed the GMP diet for 10 weeks at home indicates safety, acceptability of GMP food products, a 13-14% reduction in blood Phe levels (p<0.05) and improved distribution of dietary protein throughout the day compared with the amino acid diet. In summary, food products made with GMP that is supplemented with limiting indispensable amino acids provide a palatable alternative source of protein that may improve dietary compliance and metabolic control of PKU.

Concentrations and inactivation of Ascaris eggs and pathogen indicator organisms in wastewater stabilization pond sludge.

During treatment in wastewater stabilization ponds (WSPs) many pathogens, in particular helminth eggs, are concentrated in the sludge layer. Because periodic removal of the sludge is often required, information is needed on the concentrations and inactivation of pathogens in the sludge layer to evaluate the public health risk they pose upon removal of the sludge. In this paper, previous reports on the sludge concentrations of various pathogen indicator organisms and helminth eggs are reviewed and results from our own recent experiments are reported. The advantages and disadvantages of several methods for studying inactivation in the sludge layer are discussed, as well as implications for the management of WSP sludge. In our recent experiments, which were conducted at three WSPs in central Mexico, sludge cores, dialysis chambers, and batch experiments were used to measure the inactivation rates of fecal coliform bacteria, fecal enterococci, F+ coliphage, somatic coliphage, and Ascaris eggs. The first-order inactivation rate constants were found to be approximately 0.1, 0.1, 0.01, 0.001, and 0.001 d(-1), respectively. The concentrations of all the organisms were found to vary both vertically and horizontally in the sludge layer; therefore, to determine the maximum and average concentration of organisms in the sludge layer of a WSP, complete sludge cores must be collected from representative locations throughout the pond.

Inactivation of viable Ascaris eggs by reagents during enumeration.

Various reagents commonly used to enumerate viable helminth eggs from wastewater and sludge were evaluated for their potential to inactivate Ascaris eggs under typical laboratory conditions. Two methods were used to enumerate indigenous Ascaris eggs from sludge samples. All steps in the methods were the same except that in method I a phase extraction step with acid-alcohol (35% ethanol in 0.1 N H(2)SO(4)) and diethyl ether was used whereas in method II the extraction step was avoided by pouring the sample through a 38-microm-mesh stainless steel sieve that retained the eggs. The concentration of eggs and their viability were lower in the samples processed by method I than in the samples processed by method II by an average of 48 and 70%, respectively. A second set of experiments was performed using pure solutions of Ascaris suum eggs to elucidate the effect of the individual reagents and relevant combination of reagents on the eggs. The percentages of viable eggs in samples treated with acid-alcohol alone and in combination with diethyl ether or ethyl acetate were 52, 27, and 4%, respectively, whereas in the rest of the samples the viability was about 80%. Neither the acid nor the diethyl ether alone caused any decrease in egg viability. Thus, the observed inactivation was attributed primarily to the 35% ethanol content of the acid-alcohol solution. Inactivation of the eggs was prevented by limiting the direct exposure to the extraction reagents to 30 min and diluting the residual concentration of acid-alcohol in the sample by a factor of 100 before incubation. Also, the viability of the eggs was maintained if the acid-alcohol solution was replaced with an acetoacetic buffer. None of the reagents used for the flotation step of the sample cleaning procedure (ZnSO(4), MgSO(4), and NaCl) or during incubation (0.1 N H(2)SO(4) and 0.5% formalin) inactivated the Ascaris eggs under the conditions studied.

Chyloabdomen in a mature cat.

A mature, castrated male cat presented with progressive lethargy and a severely distended abdomen. Abdominal radiographs, abdominocentesis, and evaluation of the fluid obtained led to a diagnosis of chyloabdomen. The underlying pathology, etiology, diagnosis, and treatment associated with this disease are discussed.

Linguistic analysis to assess medically related posttraumatic stress symptoms.

The authors examined the presence of posttraumatic stress symptoms (PTSS) in 20 patients requiring ventilation after acute respiratory distress. The subjects completed a semistructured interview about their ventilation experience that was subject to content and linguistic analysis. Subjects also completed two self-report measures to assess PTSS and socioemotional adjustment. Subjects who endorsed PTSS were more likely to use a narrative style suggesting emotional involvement in their recall of the stressful event. The authors indicate that the presence of PTSS is a common consequence of traumatic medical experiences and that denial of distress may be an adaptive short-term coping strategy.

Improved detection and characterization of paroxysmal nocturnal hemoglobinuria using fluorescent aerolysin.

Paroxysmal nocturnal hemoglobinuria (PNH) is caused by a somatic mutation in the gene PIGA, which encodes an enzyme essential for the synthesis of glycosylphosphatidylinositol (GPI) anchors. The PIGA mutation results in absence or marked deficiency of more than a dozen proteins on PNH blood cells. Current flow cytometric assays for PNH rely on the use of labeled antibodies to detect deficiencies of specific GPI anchor proteins, such as CD59. However, because no single GPI anchor protein is always expressed in all cell lineages, no one monoclonal antibody can be used with confidence to diagnose PNH. We describe a new diagnostic test for PNH, based on the ability of a fluorescently labeled inactive variant of the protein aerolysin (FLAER) to bind selectively to GPI anchors. We compared GPI anchor protein expression in 8 patients with PNH using FLAER and anti-CD59. In all cases, FLAER detected similar or higher proportions of PNH monocytes and granulocytes compared with anti-CD59. Because of the increased sensitivity of detection, FLAER could detect small abnormal granulocyte populations in patients to a level of about 0.5%; samples from healthy control subjects contained substantially fewer FLAER-negative cells. FLAER gives a more accurate assessment of the GPI anchor deficit in PNH.

Channel formation by the glycosylphosphatidylinositol-anchored protein binding toxin aerolysin is not promoted by lipid rafts.

Glycosylphosphatidylinositol-anchored proteins may be concentrated in membrane microdomains (lipid rafts) that are also enriched in cholesterol and sphingolipids. The glycosyl anchor of these proteins is a specific, high affinity receptor for the channel-forming protein aerolysin. We wished to determine if the presence of rafts promotes the activity of aerolysin. Treatment of T lymphocytes with methyl-beta-cyclodextrin, which destroys lipid rafts by sequestering cholesterol, had no measurable effect on the sensitivity of the cells to aerolysin; nor did similar treatment of erythrocytes decrease the rate at which they were lysed by the toxin. We also studied the rate of aerolysin-induced channel formation in liposomes containing glycosylphosphatidylinositol-anchored placental alkaline phosphatase, which we show is a receptor for aerolysin. In liposomes containing sphingolipids as well as glycerophospholipids and cholesterol, most of the enzyme was Triton X-100-insoluble, indicating that it was localized in rafts, whereas in liposomes prepared without sphingolipids, all of the enzyme was soluble. Aerolysin was no more active against liposomes containing rafts than against those that did not. We conclude that lipid rafts do not promote channel formation by aerolysin.