Occurrence of 16s rRNA methylase ArmA producing Enterobacteriaceae in a general hospital in Warsaw, Poland

Resistance to gentamicin, amikacin and kanamycin was screened in 270 clinical isolates of Enterobacteriaceae originated from April 19 to May 19, 2010 in a regular hospital in Warsaw, Poland. Most of the isolated bacteria were considered pathogenic. Nineteen isolates (7%) were simultaneously resistant to two or three of the tested aminoglycosides. MICs of the three aminoglycosides ranged form 128 to 1024 mcg/ml for six isolates. These isolates were suspected to produce 16S rRNA methylase. Genes encoding for three methylases reported in Europe: ArmA, RmtB and RmtC were searched by PCR. The armA gene was detected in all of the six isolates. This group encompassed Enterobacter cloacae (n=4), Klebsiella pneumoniae (n=1) and Proteus mirabilis (n=1). Five isolates of this group carried the bla(CAX-M) gene for CTX-M type ESBL. The remaining isolate E. cloacae DM0340 was ESBL negative and lacked bla(CRX-M) that may suggest an altered genetic environment of the armA gene in this isolate. Our results showed that 2.2% of the tested isolates produced 16S rRNA methylase ArmA. This finding may argue for a high incidence of ArmA producing Enterobacteriaceae in Poland when compared to reports from other European countries.     

The role of Dsb proteins of Gram-negative bacteria in the process of pathogenesis

Tertiary and quaternary structures of extracytoplasmic proteins containing more than one cysteine residue often require introduction of disulfide bonds. This process takes place in an oxidative environment, such as the periplasm of Gram-negative bacteria, and is catalyzed by Dsb (disulfide bond formation) proteins. Mutations in dsb genes influence the conformation and stability of many extracytoplasmic proteins. Thus, many pathogens become partially or fully attenuated due to improper folding of proteins that act as virulence factors. This review summarizes the current knowledge on Dsb proteins and their effect on the pathogenicity of Gram-negative bacteria. The potential application of Dsb proteins in biotechnology is also discussed.     

HAX-1 protein: multifunctional factor involved in apoptosis, cell migration, endocytosis and mRNA transport

HAX-1 protein, an anti-apoptotic factor, first identified in 1997, is also involved in cell migration, endocytosis and probably mRNA transport. HAX-1 structure indicates similarity to the proteins form Bcl-2 family, although there is no strong homology. HAX-1 is a substrate for Omi/HtrA2, a protease responsible for degradation of the caspases, and functions as an inhibitor of caspase-9, which points to its role in the regulation of apoptosis. Several HAX-1 interactions with proteins involved in apoptosis and cell motility were demonstrated. Another line of inquiry focus on its ability to bind 3' untranslated regions of the certain mRNAs. Some data indicate that it might be involved in mRNA transport. HAX-1 multifunctionality and its involvement in the processes important for the cell status suggest its possible role in oncogenesis and metastasis. It is also known that HAX-1 deficiency or overexpression leads to hereditary or systemic diseases (Kostmann disease, lesional psoriasis, systemic sclerosis). Therefore, detailed analysis of HAX-1 functions could be medically important.     

The influence of glutamic and aminoacetic acids on the excitability of the liverwort Conocephalum conicum

Intracellular microelectrode measurements revealed that a resting potential (RP), an action potential (AP) and a calcium component of AP (named voltage transient, VT) can be influenced by glutamic acid (Glu) and aminoacetic acid (glycine, Gly) in the liverwort Conocephalum conicum. In the continuous presence of 5mM Glu or 5mM Gly, the RP hyperpolarized constantly and the plants became desensitized to the excitatory amino acids (Glu or Gly). Under such circumstances, the amplitudes of APs evoked by stimuli other than Glu or Gly grew, as did their calcium components (VTs). The sudden application of 1-15 mM Glu or Gly to a thallus not yet desensitized resulted in an excitation, i.e. a single AP or AP series. Aspartate (Asp) could not substitute for Glu in any way. Simultaneous action of both amino acids acted synergically to trigger APs. The same phenomenon was observed when glycine solution was enriched with N-methyl-D-aspartic acid (NMDA). Gly-induced APs were totally hindered by 1mM D-amino-5-phosphonopentanoic acid (AP5)--an inhibitor of ionotropic glutamate receptors of the NMDA kind. Glu-induced APs could be totally suppressed by 1mM AP5 as well as by 1mM 6,7-dinitroquinoxaline-2,3-dione (DNQX)--an inhibitor of AMPA/KA receptors. DNQX also completely blocked the calcium component of Glu-evoked APs. After DNQX treatment, the only response to Glu was a membrane potential hyperpolarization (like the Glu response in a desensitized plant). It was concluded that the Glu-induced depolarization and hyperpolarization are separate phenomena. The stimulatory effects of both Glu and Gly on liverwort excitability may be the consequences of an activation of a variety of ionotropic Glu receptor subtypes.     

Serum activity of dipeptidyl peptidase IV (DPPIV; EC 3.4.14.5) in breast-fed infants with symptoms of allergy

Beta-casomorphins, opioid peptides present in mother's milk, are a good substrate for DPPIV (EC 3.4.14.5) which is a major factor limiting the half-life of biologically active peptides. Serum DPPIV activity of two groups of infants (healthy and atopic dermatitis) and contents of beta-casomorphin-5 and -7 in their mothers' milk were determined in the study. We have found correlation between those two parameters in the group of children with atopic dermatitis syndromes, while no such a correlation was found in the control group.     

Changes of beta-casomorphin content in human milk during lactation

Milk is the best, complete food important for the development and nourishment of a neonate. Except for nutrients, milk contains biologically active opioid peptides derived from beta-casein, named beta-casomorphins (BCMs), which can exert effects in the gastrointestinal tract as well as in the whole body of neonates. The content of beta-casomorphins in human milk during maturation phases has not been studied so far. The aim of this study was to determine the content of beta-casomorphin-5 and -7 in human milk in different phases of lactation. A significantly higher concentration of both beta-casomorphins was found in colostrum than in mature milk. The concentration of beta-casomorphin in milk collected in the second month of lactation was similar to the level obtained in the fourth month of lactation. The content of beta-casomorphins in human milk was observed with the period of lactation. The level of opioid peptides may depend on the function of these peptides in neonate's body and may be associated with the maturation process.     

Antigens HLA-G, sHLA- G and sHLA- class I in reproductive failure

It can be supposed that relation between HLA-G polymorphism and sHLA-G protein expression are associated with successful embryo implantation and pregnancy maintenance. The aim of the study was the estimation specific differences in expression of sHLA-G and sHLA- class I antigens in women with reproductive failure in comparison with fertile women. The study sample enrolled 80 women, divided into 2 groups. The study group (B) enrolled 60 women with reproductive failure including 20 women with 3 recurrent spontaneous abortions in the first trimester of pregnancy (RSA), 20 women with empty sac (ES) and 20 women with 3 consecutive in-vitro fertilization failures (IVFf). The control group (C) enrolled 20 fertile women with at least 2 children. Soluble HLA- class I antigens (sHLA-I) and soluble HLA-G (sHLA-G) were determined using ELISA test kits from IBio Vendor Labolatory Medicine, Inc. HLA-G allele found in individuals in our study were identified by comparing the obtained bp sequences of exon 2., 3. and 4. with bp sequences of HLA-G antigen published at the Nolan Research Institute website. The highest concentration of sHLA-I is noted among women with HLA-G 10401 allele which differed significantly for the mean sHLA-I concentration calculated for all the remaining alleles (p<0.0001). The most prevalent alleles were: HLA-G 10101, 10102 and 10108 with sHLA-I concentrations among women bearing those alleles significantly lower in comparison to the HLA-G 10401 carriers (p<0.001). Allele 10101 and 10102 was related to the lower significantly plasma sHLA-I concentrations than 10108 allele (p<0.02). Lowest mean sHLA-G values were observed in the IVFf group with significant difference from the remaining groups (p<0.05). To conclude, sHLA-G molecules is associated to certain HLA-G alleles and imply that sHLA-G levels are under genetic control. Low concentration sHLA-G seems to be prognostically important in IVF failure.     

Tissue localization of tumor antigen-loaded mouse dendritic cells applied as an anti-tumor vaccine and their influence on immune response

The recognition, internalization and intracellular processing of antigen are the main functions of dendritic cells (DCs). In the course of these processes, DCs differentiate and acquire the ability to produce cytokines responsible for polarization of the immunological response. Therefore, vaccination with tumor antigen-loaded DCs is one of the most promising approaches to induce tumor-specific immune response. The purpose of this study was to analyze the migratory abilities, from an injection site to tumor-draining lymph nodes (tLN), of DCs applied as an anti-tumor vaccine and their capacity for immune response activation. Mouse DCs of the established JAWS II cell line transduced with EGFP gene or ex vivo bone marrow-isolated DCs (BM-DCs) stained with intravital CFDA dye were loaded with MC38 colon carcinoma tumor lysate (TAg) and then administered peritumorally to MC38 tumor-bearing C57BL/6 mice. On the first, third, fifth and seventh days after injection the tumors, tLNs and spleens were examined. The TAg-loaded DCs migrated more effectively to the tLNs than did the unloaded control DCs; however, the majority of them remained in the tumor vicinity. Immunohistological analysis of the tumor tissues demonstrated that only TAg-loaded DCs activated an immune response. Seven days after DCs vaccine administration, numerous necrotic areas and some apoptotic bodies were observed in the tumor tissue. However, the anti-MC38 tumor cytotoxic activity of spleen and tLN cells from mice treated with both TAg-loaded and unloaded DCs reached a maximum on the fifth day after DC injection. Concluding, TAg-loaded DCs migrated more efficiently to tLNs and were more effective activators of local (but not systemic) cellular immune response than were unloaded DCs. We hypothesize that only the application of TAg-loaded DCs to tumor-bearing mice as an adjuvant supporting chemotherapy may activate a more effective anti-tumor response.     

Assessment of prolactin secretion in children: a profile of circadian prolactin secretion and the principles for interpreting it

INTRODUCTION: Prolactin (Prl) is secreted in a circadian pattern, although no method of interpreting it has yet been established. The aim of the study was to assess Prl secretion in children on the basis of the Prl circadian profile and to establish principles for the interpretation of the results obtained by this method. MATERIAL AND METHODS: The analysis comprised 41 healthy short children (25 boys); aged 5.2-16.3 years, in whom hormonal disorders and chronic diseases had been excluded. The children were divided into prepubertal or pubertal subgroups. Serum Prl concentrations were measured every 3 hours for 24 hours. To assess the rhythm the parameters of macroscopic analysis were calculated and receiver operating characteristic (ROC) analysis was performed. The group for comparison consisted of 30 children aged 8.9-17.2 years with hyperprolactinaemia. RESULTS: In each subgroup significantly higher Prl concentrations were observed at night than by day. No statistical differences were noticed between the groups regarding Prl concentrations at particular time points or parameter values during circadian Prl rhythm evaluation. In the group analysed weak correlations were found between age and Prl peak and trough levels. On the basis of ROC analysis criteria for the existence of normal circadian Prl rhythm in children were established. CONCLUSIONS: 1. The presence of normal circadian Prl rhythm is observed if at least one of the following three criteria is fulfilled: amplitude >1.8779; X(n)/X(d) ratio >1.685; regression index <-0.4107. 2. No interpretation in relation to sex, age and stage of puberty is necessary for the circadian prolactin secretion rhythm in children.