Early life adversity and telomere length: a meta-analysis.

Early adversity, in the form of abuse, neglect, socioeconomic status and other adverse experiences, is associated with poor physical and mental health outcomes. To understand the biologic mechanisms underlying these associations, studies have evaluated the relationship between early adversity and telomere length, a marker of cellular senescence. Such results have varied in regard to the size and significance of this relationship. Using meta-analytic techniques, we aimed to clarify the relationship between early adversity and telomere length while exploring factors affecting the association, including adversity type, timing and study design. A comprehensive search in July 2016 of PubMed/MEDLINE, PsycINFO and Web of Science identified 2462 studies. Multiple reviewers appraised studies for inclusion or exclusion using a priori criteria; 3.9% met inclusion criteria. Data were extracted into a structured form; the Newcastle-Ottawa Scale assessed study quality, validity and bias. Forty-one studies (N=30 773) met inclusion criteria. Early adversity and telomere length were significantly associated (Cohen's d effect size=-0.35; 95% CI, -0.46 to -0.24; P<0.0001). Sensitivity analyses revealed no outlier effects. Adversity type and timing significantly impacted the association with telomere length (P<0.0001 and P=0.0025, respectively). Subgroup and meta-regression analyses revealed that medication use, medical or psychiatric conditions, case-control vs longitudinal study design, methodological factors, age and smoking significantly affected the relationship. Comprehensive evaluations of adversity demonstrated more extensive telomere length changes. These results suggest that early adversity may have long-lasting physiological consequences contributing to disease risk and biological aging.

Identification of human rotavirus strains with the P[14] genotype by PCR.

A seminested PCR typing assay has been extended to identify rotavirus strains with the P[14] genotype. The specificity of the method was confirmed by Southern hybridization and by restriction analysis with the enzyme AluI. One out of four human rotavirus (HRV) strains with unusual subgroup-electropherotype linkage but none out of 50 HRV strains with usual linkage was typed as P[14].

[Clinical comparison of different replacement milk formulas in children with allergies to cow's milk proteins. 24-month follow-up study]. / Comparazione clinica di differenti formule di latte sostitutivo in bambini con allergia alle proteine del latte vaccino. Follow-up a 24 mesi.

The elective treatment for allergy to cow's milk protein is the elimination of these proteins from the diet. The present study with a follow-up of over two years took the form of a comparison between different replacement formulas based on soya (group A), hydrolysate of soya and bovine collagen (group B), and hydrolysate of casein (group C), randomly administered to 55 children (30 males and 25 females, aged between 2-48 months) with documented allergy to cow's milk proteins, but with different clinical symptoms. Tests to evaluate the acquisition of clinical tolerance to cow's milk proteins were performed using a day-hospital regime every 6 months. Sensitivity reactions were observed in 22% of cases in group A, 8% in group B and 37.5% in group C. It is worth underlining that 5 of the 6 children with reactions to soya protein then showed an excellent tolerance to hydrolysate of soya when it was administered subsequently until tolerance was achieved. Weight and statutory growth was uniformly good in all three groups. A high percentage of children achieved tolerance after 24 months (72%); the mean time taken to acquire clinical tolerance was 11.6 +/- 4.8 in group A, 11.6 +/- 6.02 in group B, and 14 +/- 5.6 in group C. No correlation was found between the type of initial symptoms, age at onset, method of response to first challenge and the time taken to acquire tolerance.

Trisomy 9 mosaicism syndrome. A case report and review of the literature.

The authors report on a case of trisomy 9 mosaicism syndrome, a rare chromosome abnormality. The common features of this syndrome are growth and mental retardation, low-set malformed ears, wide sutures and fontanelles, bulbous nose, short palpebral fissures, micrognathia, microphthalmia and enophthalmos, abnormal hands and feet, hip dislocation, joint limitation, cardiovascular defects and urogenital abnormalities. Our patient presented some unusual characteristics, such as 13 pairs of ribs, a vertebral malformation, a hemivertebra and a Dandy-Walker syndrome. They compare their clinical findings with the few cases previously described and they try to contribute to the further clinical definition of the syndrome. It is possible that there is a correlation between the variability of the phenotype and the percentage of trisomic cells in the patient.

Management of incipient caries in young adults. Dentists state their preference.

Survey finds general dentists in upstate New York use variety of treatment modalities to manage questionable or early caries in their young adult patients. The age of the dentist appears to be a factor in treatment selection, as is location of practice.

Production of analytical quantities of recombinant proteins in Chinese hamster ovary cells using sodium butyrate to elevate gene expression.

Sodium butyrate was used to enhance expression levels and thereby facilitate the generation of analytical quantities of nine different tissue plasminogen activator (tPA) analogues expressed under the control of the cytomegalovirus immediate early (CMV IE) promoter by the Chinese hamster ovary (CHO) mammalian expression system. Production involved growth in roller bottles, using serum free or low serum media formulations, together with repetitive, sodium butyrate inductions. Average inductions in the presence of sodium butyrate ranged from 2 to 9-fold relative to uninduced controls, using cell lines with no previous butyrate exposure. Retardation of growth rate by butyrate minimized the need to split cells during the production runs, extending longevity of roller bottles containing cells secreting at induced levels. SDS-PAGE analyses indicate a consistently high percentage of single-chain material. Measurements of specific activity and fibrinogen fragment enhancement for one of the analogues demonstrate that neither of these two critical parameters are affected by production in the presence of butyrate. Induction kinetic data and growth curves for the expression of sCD4 under control of the SV40 early promoter demonstrate that the benefits of butyrate can be realized with different promoters and heterologous genes, and are additive when used in conjunction with an amplified cell line constitutively expressing at elevated levels. This work demonstrates the practical application of sodium butyrate in the production of analytical quantities of protein from the CHO expression system, and suggests a role for sodium butyrate in commercial scale processes as well.

Construction of a tissue plasminogen activator gene having unique restriction sites to facilitate domain manipulation: a model for the analysis of multi-domain proteins.

We have designed and constructed a DNA sequence encoding human tissue plasminogen activator (tPA) with convenient restriction sites that flank each of the domains of the heavy chain. To accomplish this, the first 1095 bases of the gene coding for the mature protein were synthesized with unique restriction sites engineered into the interdomainal regions. This synthetic construction was then ligated to a cDNA fragment of the tPA gene that encoded the active site, thus generating a full-length tPA gene. The gene products produced by Chinese hamster ovary (CHO) cells transfected with either the tPA cassette gene or the tPA cDNA gene were then compared with the tPA produced by Bowes melanoma cells to determine whether or not synthetic interdomainal amino acid changes had an effect on the biochemical characteristics of the molecule. Specifically, molecular weight, specific activity, enhancement by fibrinogen fragments and kinetic constants were analysed. None of the properties examined were significantly different from those of the native melanoma tPA. Therefore, the cassette gene described herein should provide considerable versatility and precision in the construction of tPA mutants by facilitating the manipulation of the finger, growth factor and kringle domains, and likewise should be useful in assessing the function of these domains within the tPA molecule. We present this cassette gene system as a model for the analysis of protein domain function applicable to other multi-domain proteins.

Knowledge and the child's developing theory of the world.

In this article, I have presented a brief history of research directed toward examining the developing ability of children to comprehend and produce metaphor that occurs with increasing age. I argued that the child's ability in this area does not rest upon developing an ability specific to metaphor, but, instead, metaphor comprehension and production, not unlike other forms of cognition, rests upon a developing theory of the world founded upon an expanding knowledge base. Given that knowledge is central to the child's cognitive development, I turned to an examination of the nature of knowledge. The discussion of knowledge drawn from philosophers who have attempted to propose theories of knowledge in answer to the question, "What is knowledge?" Traditional justificationalist theories of knowledge and subsequently developed nonjustificationalist theories were considered. The discussion was focused upon a number of issues which have divided philosophers with respect to their treatment of knowledge in order to highlight the problematic nature of knowledge. Cognitive and developmental psychologists may wish to consider some of these issues and the implications they hold for their theories as they study knowledge and/or invoke knowledge as a part of their theory construction. Some of the implications of the issues were illustrated in a discussion of the relation of knowledge to the developing child's comprehension and production of metaphor.

Localization of thymostimulin in mammalian thymuses: comparative evaluation.

An anti-thymostimulin (TS) serum was tested on rabbit, guinea-pig, lamb and pig thymuses to study the localization of the hormonal factor. The immuno-peroxidase method, with some modifications, was applied to tissue fixed in Bouin's or in Bouin-PAF liquids and embedded in paraffin wax. Immuno-reactivity was shown in reticulo-epithelial cells in the cortex, in the medulla and in the external cells of the Hassall's corpuscles. The intensity of the immuno-reaction is stronger in pig thymus, whereas in lamb, rabbit and guinea-pig thymuses it is more uniform and pale, but to varying degrees. Comparison with the localization of TS in thymuses of other mammals and man and the problem of species-specificity were discussed.

Laboratory studies of acute varicella and varicella immune status.

We evaluated varicella-zoster virus (VZV) culture and serum antibody methods utilizing specimens from 620 children enrolled in protocols for prevention or treatment of varicella and samples routinely submitted to the Clinical Virology Laboratory. In a foreskin fibroblast tube culture system, we initially isolated VZV from only 29 (51%) of 57 children cultured on the first day of varicella. After modifying the method, the proportion of culture-positive children increased significantly to 36 (80%) of 45 (p less than 0.005 by corrected X2), and the median days-to-positivity were significantly shortened from 5.6 to 3.8 days (p less than 0.001, Wilcoxon rank-sum test). The Viran fluorescent antibody to membrane antigen (FAMA) assay was difficult to read and not reproducible. The standard FAMA was more sensitive than the Merck ELISA antibody test for detecting vaccine-induced antibody. The Whittaker ELISA did not detect vaccine-induced antibody but was comparable to FAMA for immune status testing (sensitivity, 95%; specificity, 92%) and for diagnosis of acute varicella.

Expression of a cloned lipopolysaccharide antigen from Neisseria gonorrhoeae on the surface of Escherichia coli K-12.

A gonococcal gene bank maintained in Escherichia coli K-12 was screened by colony immunoblotting, and a transformant expressing a surface antigen reactive to anti-gonococcal outer membrane antiserum was isolated. The isolate carried a recombinant plasmid, pTME6, consisting of approximately 9 kilobases of Neisseria gonorrhoeae DNA inserted into the BamHI site of pBR322. Surface labeling of E. coli HB101(pTME6) confirmed that the antigen was expressed on the E. coli cell surface. The antigenic material was resistant to proteinase K digestion and sensitive to periodate oxidation, indicating that the material was carbohydrate. Purified lipopolysaccharide (LPS) from HB101(pTME6) produced a unique band on silver-stained polyacrylamide gels that contained immunoreactive material as seen on Western blots of LPS samples. Only two of three E. coli LPS mutant strains carrying pTME6 reacted with the antigonococcal antiserum, suggesting that a certain E. coli core structure is necessary for antigen expression. We conclude that pTME6 contains one or more gonococcal genes encoding an LPS core biosynthetic enzyme(s) which can modify E. coli core LPS to produce a gonococcuslike epitope(s).

Use of lambda exonuclease for efficient oligonucleotide-mediated site-directed deletion and point mutation of double-stranded DNA.

A novel approach to oligonucleotide-mediated, site-directed in vitro mutagenesis is described that allows for the efficient generation of sequence modifications on double-stranded substrates without the need for subcloning into special vectors. Site-directed deletions as well as point mutations were introduced into the genes encoding human tissue plasminogen activator (tPA) and the Bacillus amyloliquefaciens alpha-amylase gene using lambda exonuclease to enzymatically degrade DNA 5' to 3' in order to generate a single-stranded template in the immediate vicinity of the oligonucleotide annealing site. The mutagenizing oligonucleotide, used both to redefine the 5' end of the molecule and to introduce base changes, was annealed to the single-stranded target sequence producing substrates for both the exonucleolytic and polymerizing activities of DNA polymerase Klenow fragment. Resolution of the resultant heteroduplex by Escherichia coli resulted in the generation of the desired deletion point mutation in the tPA sequence with an efficiency of 38% as determined by differential hybridization and 32% as determined by restriction analysis, with final verification by sequence data. As a further test of the method, two point mutations were introduced simultaneously with the desired sequence deletion into the Bacillus amyloliquefaciens alpha-amylase gene, generating a Pst I restriction site at the junction of the DNA encoding the signal peptide and the mature enzyme with an efficiency of 0.3% as determined by sequence data of hybridization-positive/Pst I-positive clones. The lambda exonuclease procedure is designed for use in situations where site-directed deletions must be introduced efficiently alone or with single or double point mutations.

Induction and repression of outer membrane proteins by anaerobic growth of Neisseria gonorrhoeae.

Neisseria gonorrhoeae is generally considered to be an obligate aerobe; it can, however, grow in the absence of oxygen by anaerobic respiration by using nitrite as a terminal electron acceptor. The outer membrane protein compositions of aerobically and anaerobically grown N. gonorrhoeae strains were compared by one- and two-dimensional polyacrylamide gel electrophoresis. Anaerobically grown strains expressed at least three proteins (Pan 1 to Pan 3) at much higher levels than did aerobically grown cells. Conversely, at least five other proteins (Pox 1 to Pox 5) were found to be expressed at significantly higher levels in aerobically grown cells. None of the Pan or Pox proteins were heat modifiable, and none of the heat-modifiable protein IIs or other major outer membrane proteins (protein I, protein III, pilin, or H-8 protein) were significantly altered in expression by anaerobic growth. There were also no apparent differences in lipopolysaccharide composition in aerobically and anaerobically grown gonococci. The regulation of protein expression by oxygen availability suggests that anaerobic growth is a physiologically significant state for this organism.

Pseudorabies virus as a live virus vector for expression of foreign genes.

The cDNA coding for human tissue plasminogen activator (tPA) was cloned downstream from the promoter for pseudorabies virus (PRV) glycoprotein and flanked by downstream PRV DNA. After co-transfection with PRV DNA, this plasmid recombined to insert the tPA cDNA into the viral genome. In cells infected by this recombinant virus, tPA was detected by immunoprecipitation analysis and by enzymatic activity. Since it has a wide host range but does not infect humans, PRV is a possible vaccine vector for genes from animal pathogens.

Isolation and characterization of native human renin derived from Chinese hamster ovary cells.

Transfection of Chinese hamster ovary (CHO) cells with a plasmid containing the cDNA for human preprorenin has provided cell lines that secrete 15 mg of native prorenin per liter of culture medium. Tryptic activation of the prorenin occurs by selective cleavage of the Arg66-Leu67 bond (numbering as in preprorenin). The renin product, purified in a single step and in high yield by affinity chromatography, is fully stable for as long as 8 months when stored in solution at 4 degrees C and pH 6.5. Purity of the renin was judged to be greater than 95% by gel electrophoresis, compositional and N-terminal sequence analyses, and specific enzyme activity. An important aspect of the present work is the development of a direct assay for renin which permits accurate and reproducible evaluation of enzyme units and kinetic parameters. Application of methods described herein, combined with appropriate scale-up fermentation capabilities, provides the means for generating gram quantities of human renin and its zymogen.

Localization of thymostimulin in mouse and rat thymuses.

Anti-bovine thymostimulin serum reacts with reticulo-epithelial cells in the cortex and in the medulla of rat and mouse thymuses. The immuno-reaction is present in a smaller number of cells which have more pallid color than those found in calf thymus. Comparative examination as to the distribution of other thymic factors and as to species-specificity was conducted.