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Objective: To determine the frequency of mitochondrial DNA depletion syndrome (MDS) in infants with cholestasis and liver failure and to further clarify the clinical, biochemical, radiologic, histopathologic, and molecular features associated with MDS due to deoxyguanosine kinase (DGUOK) and MPV17 gene mutations., Study Design: We studied 20 infants with suspected hepatocerebral MDS referred to our tertiary care center between 2007 and 2013. Genomic DNA was isolated from blood leukocytes, liver, and/or skeletal muscle samples by standard methods. Mitochondrial DNA copy number relative to nuclear DNA levels was determined in muscle and/or liver DNA using real-time quantitative polymerase chain reaction and compared with age-matched controls. Nuclear candidate genes, including polymerase γ, MPV17, and DGUOK were sequenced using standard analyses., Results: We identified pathogenic MPV17 and DGUOK mutations in 11 infants (6 females) representing 2.5% of the 450 cases of infantile cholestasis and 22% of the 50 cases of infantile liver failure referred to our center during the study period. All of the 11 patients manifested cholestasis that was followed by a rapidly progressive liver failure and death before 2 years of life. Mitochondrial DNA depletion was demonstrated in liver or muscle for 8 out of the 11 cases where tissue was available. Seven patients had mutations in the MPV17 gene (3 novel mutations), 4 patients had DGUOK mutations (of which 2 were novel mutations)., Conclusion: Mutations in the MPV17 and DGUOK genes are present in a significant percentage of infants with liver failure and are associated with poor prognosis., (Copyright © 2014 Mosby, Inc. All rights reserved.)

Objective: To compare the performance of five risk models (Diamond-Forrester, the updated Diamond-Forrester, Morise, Duke, and a new model designated COronary Risk SCORE (CORSCORE) in predicting significant coronary artery disease (CAD) in patients with chest pain suggestive of stable angina pectoris., Methods: Retrospective cohort for creation of CORSCORE by means of logistic regression analysis. Prospective cohort for validation of the five risk models using receiver operating characteristics (ROC) curve analysis, net reclassification improvement (NRI), and integrated discrimination improvement (IDI). Significant CAD was defined as lumen area diameter reduction ≥50% at coronary angiography. All risk models include information on age, sex, and symptoms. In addition the Duke, Morise, and CORSCORE models include information on tobacco use and hypercholesterolemia. Duke and Morise also include information on diabetes. History of myocardial infarction is used by the Duke and CORSCORE models whereas hypertension is included in the Morise and CORSCORE models. The Duke model includes information on electrocardiogram (ECG) changes and the Morise model includes information on family history, body mass index, obesity, and oestrogen status., Results: 4781 retrospective and 633 prospective consecutive patients referred for coronary angiography were included. The area under the ROC for the updated Diamond-Forrester, Duke, and CORSCORE was significantly larger than for the Diamond-Forrester (p≤0.001). The IDI was significantly higher for the Duke as compared to all other models (p≤0.006)., Conclusion: The Duke, updated Diamond-Forrester, and CORSCORE risk models are most efficient in predicting CAD in a contemporary cohort of patients with symptoms suggestive of angina. The updated Diamond-Forrester may most operational in daily clinical practice since it is calculated from the lowest number of clinical variables., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Recombinant polyclonal antibodies are a new class of protein biologics, combining a defined number of target-specific antibodies, developed for therapeutic use across various indications. Development, manufacture, and release of recombinant polyclonal antibodies as well characterized biological products have required development of new chemistry, manufacturing, and control (CMC) technologies. Sym001 is a recombinant polyclonal antibody product containing 25 unique antibodies specific for the Rhesus D antigen. Sym001 drug substance is manufactured using a single batch technology, Sympress. Here, we describe the development of two novel mass spectrometry based methods that allows identification of individual antibodies in the Sym001 drug substance, through the determination of unique marker peptides or antibody light chains. The two methods provide an unambiguous identification of the 25 unique antibodies comprised in the Sym001 drug substance. Furthermore, the light chain liquid chromatography-mass spectrometry (LC-MS) method has been developed to allow the determination of the relative distribution of the 25 antibodies. The light chain LC-MS method has demonstrated linearity, specificity, precision, and accuracy, thus qualifying it for use in the quality control of recombinant polyclonal antibodies for human use. The development of such quantitative methods is central for the development and quality control of additional therapeutic recombinant polyclonal antibody products.

Objective: This study was undertaken to evaluate the risk of persistent trophoblastic disease and obstetric complications related to a multiple pregnancy comprising a diploid hydatidiform mole and normal cofetus(es)., Study Design: From a database of 270 consecutively collected hydatidiform moles, 8 multiple and 154 singleton molar pregnancies were identified. Molar and fetal ploidy was determined, and data on clinical features and chemotherapy were collected. Differences between groups were assessed with Fisher's exact or Mann-Whitney test., Results: The molar component in all 8 multiple pregnancies was diploid. Five patients with diploid mole and coexisting fetus pregnancy chose to terminate their pregnancy, 2 aborted spontaneously, and 1 patient delivered a healthy child. Two diploid mole and coexisting fetus pregnancies (25%) and 17% of the singleton molar pregnancies were followed by persistent trophoblastic disease (P = .63)., Conclusion: The risk of persistent trophoblastic disease after a diploid mole with coexisting fetus pregnancy is similar to that after a singleton molar pregnancy, and expectant management instead of therapeutic abortion can be pursued.

Purpose: To describe a case of lightning injury restricted to the eyes and facial skin., Methods: Case history describing the clinical examination of a 54-year-old woman., Results: Following a lightning stroke the patient suffered from a sharply demarcated facial skin burn and bilateral iridocyclitis with raised intraocular pressure. Initially she had amnesia regarding the incident. She recovered on symptomatic treatment, with dry eyes as the only sequela., Conclusion: We suggest that a lightning current travelled over the outside of the patient's body facilitated by her wet raincoat, a so-called flash-over. Thereby, she was spared from more severe injury, and only the exposed areas of the face and eyes were affected.

Caseins are highly phosphorylated milk proteins assembled in large colloidal structures termed micelles. In the milk of ruminants, alphas1-casein has been shown to be extensively phosphorylated. In this report we have determined the phosphorylation pattern of human alphas1-casein by a combination of matrix-assisted laser desorption mass spectrometry and amino acid sequence analysis. Three phosphorylation variants were identified. A nonphosphorylated form, a variant phosphorylated at Ser18 and a variant phosphorylated at Ser18 and Ser26. Both phosphorylation sites are located in the amino acid recognition sequence of the mammary gland casein kinase. Notably, no phosphorylations were observed in the conserved region covering residues Ser70-Glu78, which is extensively phosphorylated in the ruminant alphas1-caseins.

Interferons (IFNs) are a family of cytokines with growth inhibitory, and antiviral functions. IFNs exert their biological actions through the expression of more than 1000 IFN stimulated genes, ISGs. ISG12 is an IFN type I induced gene encoding a protein of M(r) 12,000. We have identified a novel, IFN inducible splice variant of ISG12 lacking exon 2 leading to a putative truncated protein isoform of M(r) 7400, ISG12-S. In cells from blood and cervical cytobrush material from healthy women, the level of ISG12-S expression was higher than ISG12 expression, whereas the expression pattern was more evenly distributed between ISG12 and ISG12-S in breast carcinoma cells, in cancer cell lines and in cervical cytobrush material with neoplastic lesions. In addition, we have found a nine-nucleotide deletion situated in exon 4 of the ISG12 gene. This deletion leads to a three-amino-acid deletion (AMA) in the putative ISG12 gene products, ISG12Delta and ISG12-SDelta. We have determined the prevalence of the deletion ISG12Delta in normal and neoplastic cells. Homozygosity ISG12(0/0) and ISG12(Delta/Delta), and heterozygosity ISG12(0/Delta) were found, although the ISG12(Delta/Delta) genotype was rare. In heterozygous cells from cytobrush material with neoplastic lesions, we found a preference for expression of the ISG12(0) allele.

The present work reports isolation and characterization of a highly glycosylated protein from bovine milk fat globule membranes, known as PAS III. Partial amino-acid sequencing of the purified protein allowed construction of degenerate oligonucleotide primers, enabling isolation of a full-length cDNA encoding a protein of 330 amino-acid residues. N-terminal amino-acid sequencing of derived peptides and the purified protein confirmed 76% of the sequence and demonstrated presence of a cleavable signal peptide of 23 residues, leaving a mature protein of 307 amino acids. Database searches showed no homology to any other proteins. A survey of the human genome indicated the presence of a corresponding gene on chromosome band 11p14.3. Isolation and sequencing of the complete cDNA sequence of the human homologue proved the existence of the gene product (334 amino-acid residues). This novel mucin-like protein was named MUC15 by appointment of the HUGO Gene Nomenclature Committee. The deduced amino-acid sequences of human and bovine MUC15 demonstrated structural hallmarks characteristic for other membrane-bound mucins, such as a serine, threonine, and proline-rich extracellular region with several potential glycosylation sites, a putative transmembrane domain, and a short cytoplasmic C-terminal. We have shown the presence of O-glycosylations, identified N-glycosylations at 11 of 15 potential sites in bovine MUC15, and a splice variant encoding a short secreted mucin. Finally, analysis of human and bovine cDNA panels and libraries showed MUC15 gene expression in adult human spleen, thymus, prostate, testis, ovary, small intestine, colon, peripheral blood leukocyte, bone marrow, lymph node, tonsil, breast, fetal liver, bovine lymph nodes and lungs of both species.

Cellular growth responses of rainbow trout (Salmo gairdneri) fed different levels of dietary protein (35, 45, and 55%), with and without the anabolic steroid ethylestrenol, have been studied over a 60-day period. With an increase in dietary protein, total liver proteins increased in fish fed the steroid-free (control) diets, whereas no change occurred in the other tissues. Muscle and spleen RNA was unchanged, but RNA increased in liver, kidney, and brain. The DNA content increased in muscle, decreased in brain, but remained constant in liver, kidney, and spleen. Feeding the ethylestrenol-supplemented (experimental) diets resulted in an increase in total proteins, RNA, and DNA of kidney over the respective control value at each level of dietary protein. In the other tissues, total proteins and DNA were essentially unchanged, but total RNA content decreased in liver and increased in muscle in the experimental groups. It is concluded that in trout, the dietary protein level exerts marked differential effects on cellular growth parameters (RNA/DNA, RNA/protein, protein/DNA), which are further modified by steroid treatment. Evidence that cellular growth responses in muscle keep pace with total body growth was also indicated.

Two compounds, 3-hydroxy-3-acetonyl-oxindole (I) and compound 3-acetonylidene-oxindole (II) were synthesized by a convenient method, and their pharmacological/toxicological effects were studied in rabbit. These compounds produced transitory effects on serum enzymes. The cholesterol mobilizing effect is more prominent with compound (I).

In order to determine if the inability of thyroxine to induce cellular effects at low temperature is mediated through a temperature-sensitive system for the translocation of T4 into the nucleus, the effect of temperature on the uptake of T4 by body tissues and sub-cellular fractions of carp liver and muscle was studied in vivo. A single injection of 125I-T4 (1 micro C/10 g body weight) was given intraperitoneally to juvenile carp maintained at 15 and 25 C. Uptake from the peritoneal cavity was rapid. All the tissues exhibited maximum radioactivity at 2 hour after the injection. Fish kept at 25 C showed another peak at 8 hour and those at 15 C at 48 hour after the single injection. Transfer of T4 from the cytoplasm to nuclei was not blocked at lower temperatures. For example, in liver at 8 hour, nuclei from fish tissues kept at lower or higher temperatures had equal amounts of radioactivity. Muscle nuclei had 15% more radioactivity than liver nuclei when expressed as radioactivity/g tissue. Since there are comparable amounts of activity in the nuclei at both temperatures, some other mechanism/s than a simple block in transport from cytoplasm to nuclei is operating. There are some indications that nutritional status of fish may be playing some role in this respect.

A study on the effect of three different anabolic-androgenic steroids on the growth, food conversion efficiency and nucleic acid contents of liver, kidney, brain and muscle of carp,Cyprinus carpio was undertaken. The three steroids, methyltestosterone (MT), ethylestrenol (EE) and oxandrolone (ON), were fed in different combinations at final concentrations of 5 or 6 mg/kg diet for 60 days. No effect on the growth was observed in any of the experimental groups. A decrease in the specific growth rate (11-21%) and food conversion efficiency (20-29%) was noted. Feeding of drugs increased the cranio-somatic and reno-somatic index in all except one group. Hepatosomatic (ON+MT) and viscero-somatic (ON+MT; EE+MT+ON) indices decreased. Protein increased and RNA/DNA decreased in only one group while a decrease in protein/DNA was observed in the liver of all experimental groups. RNA/DNA increased and protein/RNA decreased only in one group while no effect was seen in protein and protein/DNA contents in any of the treated kidneys. Proteins, protein/RNA and protein/DNA decreased in certain groups in brain tissue. In muscle, no effect was seen in proteins or protein/DNA. Protein/RNA decreased in all but one group while RNA/DNA was higher only in the group fed all the three steroids together.