Molecular characterization of a patient with del(1)(q23–q25)

Summary We report a patient (S.T.) with multiple congenital anomalies and developmental delay associated with an interstitial deletion of 1q23–1q25. Molecular analysis of the deletion was performed using DNA markers that map to 1q. Five DNA markers, MLAJ-1 (D1S61), CRI-L1054 (D1S42), HBI40 (D1S66), OS-6 (D1S75), and BH516 (D1S110), were demonstrated to be deleted. Informative polymorphisms demonstrated this to be a de novo deletion of the maternally derived chromosome. Deletion status was determined using restriction fragment length polymorphism (RFLP) analysis supplemented with densitometry in the experiments where RFLP analysis was not fully informative. Deletions were confirmed by Southern analysis using genomic DNA from a somatic cell hybrid retaining the del(1)(q23–q25) chromosome that was constructed from patient S.T. Flow karyotyping confirmed the deletion and estimated that the deletion encompassed 11,000–16,000 kb. The clinical and cytogenetic characteristics of S.T. are compared with those of ten previously described patients with monosomy 1q21–1q25.     

Pig kidney dopa decarboxylase: inactivation by iodoacetamide and sequence of the carboxyamidomethylcysteine-containing peptide

Pig kidney 3,4-dihydroxyphenylalanine (Dopa) decarboxylase is inactivated by iodoacetamide following pseudo-first order reaction kinetics. The apparent first order rate constant for inactivation is proportional to the concentration of iodoacetamide and a second order rate constant of 37 M-1 min-1 is obtained at pH 6.8 and 25 degrees C. Cyanogen bromide fragmentation of iodo(1-14C)acetamide - modified inactivated Dopa decarboxylase followed by trypsin digestion yields a single radioactive peptide. Automated Edman degradation reveals a heptapeptide sequence which contains labeled carboxyamidomethylcysteine. This finding and the results of the incorporation of the label from ido (1-14C)acetamide into the enzyme clearly indicate that the modification of 1 mol of SH per mol of enzyme dimer is responsible for the inactivation process. The labeled peptide, which was located by means of limited proteolysis on the fragment corresponding to the COOH-terminal third of the enzyme, has been aligned with a 7 amino acid stretch of Drosophila enzyme. Although this region appears highly conserved in the Dopa decarboxylase enzymes, the cysteinyl residue is not conserved. This observation together with the spectral binding properties of the iodoacetamide inactivated enzyme argue against a functional role for the modifiable cysteine in the mechanism of action of pig kidney enzyme. It is suggested that the loss of pig kidney decarboxylase activity produced by iodoacetamide modification might be attributable to steric hindrance. This could be due to the presence of the bulky acetamidic group on a cysteine residue at, or near, the active center or in a site of strategic importance to the maintenance of the active site topography.     

Tyrosinase Inhibition: General and Applied Aspects

The active site of tyrosinase is described with a view to depicting its interactions with substrates and inhibitors. Occurrence and mechanism(s) of tyrosinase-mediated browning of agrofood products are reviewed, with regard to both enzymic and chemical reactions, and their control, modulation, and inhibition. Technical and applicational implications are discussed.     

Carvedilol Prevents Ovariectomy-Induced Myocardial Contractile Dysfunction in Female Rat

Carvedilol has beneficial effects on cardiac function in patients with heart failure but its effect on ovariectomy-induced myocardial contractile dysfunction remains unclear. Estrogen deficiency induces myocardial contractile dysfunction and increases cardiovascular disease risk in postmenopausal women. Our aim was to investigate whether carvedilol, a beta receptor blocker, would prevent ovariectomy-induced myocardial contractile dysfunction. Female rats (8 weeks old) that underwent bilateral ovariectomy were randomly assigned to receive daily treatment with carvedilol (OVX+CAR, 20 mg/kg), placebo (OVX) and SHAM for 58 days. Left ventricle papillary muscle was mounted for isometric tension recordings. The inotropic response to Ca²⁺ (0.62 to 3.75 mM) and isoproterenol (Iso 10⁻⁸ to 10⁻² M) were assessed. Expression of calcium handling proteins was measured by western blot analysis. Carvedilol treatment in the OVX animals: prevented weight gain and slight hypertrophy, restored the reduced positive inotropic responses to Ca²⁺ and isoproterenol, prevented the reduction in SERCA2a expression, abolished the increase in superoxide anion production, normalized the increase in p22^phox expression, and decreased serum angiotensin converting enzyme (ACE) activity. This study demonstrated that myocardial contractile dysfunction and SERCA2a down regulation were prevented by carvedilol treatment. Superoxide anion production and NADPH oxidase seem to be involved in this response.     

Mass spectrometry applications in microbiology beyond microbe identification: progress and potential

Introduction: Mass spectrometry (MS), particularly MALDI-time of flight (MALDI-TOF), has become a routine tool for microorganism identification in clinical microbiology laboratories in the last five years. The use of MALDI-TOF MS has accelerated laboratory analysis, thus providing accurate species-level information with very short turnaround times.

Areas covered: Beyond microbe identification, MALDI-TOF MS offers great opportunities for fast strain typing and detection of antimicrobial susceptibility/resistance in both bacterial and fungal organisms. Drawing on evidence from PubMed literature searches, clinical microbiology laboratory experience, and the authors’ opinions, this review summarizes recent significant advances and ongoing challenges in these areas.

Expert commentary: In the near future, it is expected that the implementation of new analytical algorithms, automation of procedures, and refinement of assays will enhance the clinical and epidemiological usefulness of MALDI-TOF and other MS technologies.     

MicroRNAs expression profiles as diagnostic biomarkers of gastric cancer: a systematic literature review

Objective: The early identification of gastric cancer (GC) represents a major clinical challenge. We conducted a systematic review of studies evaluating the miRNA expression profiling as a diagnostic tool in GC.

Methods: We performed a search of PubMed, ISI Web of Science and SCOPUS databases for studies on diagnostic miRNAs and GC, published in English up to October 2017. Eligibility criteria included case-control studies evaluating blood or tissue-based miRNA expression profiles, and incorporating at least two detection phases (screening and validation).

Results: We included 27 eligible studies, that reported on 97 deregulated miRNAs either in blood or tissue, out of which 30 were reported in at least two studies. Among 22 studies on tissue-diagnostic miRNAs, 13 consistently upregulated miRNAs (miR-214, miR-21, miR-103, miR-107, miR-196a, miR-196b, miR-7, miR-135b, miR-222, miR-23b, miR-25, miR-92 and miR-93), and six consistently downregulated miRNAs (miR-148a, miR-375, miR-133b, miR-30a, miR-193a and miR-204) were reported. Ten miRNAs with inconsistent direction of expression in tissues were identified. Among the five studies performed on blood samples, only one miRNA was consistently upregulated (miR-20a).

Conclusions: This review shows that some tissue or blood miRNAs may be considered as potential biomarkers for GC diagnosis, that urgently needs to be confirmed from large prospective studies.     

Tyrosinase inhibition: general and applied aspects

The active site of tyrosinase is described with a view to depicting its interactions with substrates and inhibitors. Occurrence and mechanism(s) of tyrosinase-mediated browning of agrofood products are reviewed, with regard to both enzymic and chemical reactions, and their control, modulation, and inhibition. Technical and applicational implications are discussed.     

Characterization of the OFD1/Ofd1 genes on the human and mouse sex chromosomes and exclusion of Ofd1 for the Xpl mouse mutant

Oral-facial-digital type 1 (OFD1) syndrome is an X-linked dominant condition characterized by malformations of the face, oral cavity, and digits. The responsible gene, OFD1, maps to human Xp22 and has an unknown function. We isolated and characterized the mouse Ofd1 gene and showed that it is subject to X-inactivation, in contrast to the human gene. Furthermore, we excluded a role for Ofd1 in the pathogenesis of the spontaneous mouse mutant Xpl, which had been proposed as a mouse model for this condition. Comparative sequence analysis demonstrated that OFD1 is conserved among vertebrates and absent in invertebrates. This analysis allowed the identification of evolutionarily conserved domains in the protein. Finally, we report the identification of 18 apparently nonfunctional OFD1 copies, organized in repeat units on the human Y chromosome. These degenerate OFD1-Y genes probably derived from the ancestral Y homologue of the X-linked gene. The high level of sequence identity among the different units suggests that duplication events have recently occurred during evolution.