Cerebrovascular response in infants and young children following severe traumatic brain injury: a preliminary report

PURPOSE: Endotoxin (lipopolysaccharide [LPS])-induced systemic organ injury leads to disruption of normal systemic organ metabolic processes, which are manifest clinically by signs of accelerated anaerobic metabolism (e.g., tissue acidosis and hyperlactatemia) and altered VO2-DO2 relationships. The association of increased anaerobic metabolism with VO2-DO2 alterations has led to the notion that ischemia/ reperfusion (I/R) injury may be a prerequisite for the development of VO2-DO2 alterations during endotoxemia. However, in contrast to sepsis, in which oxygen consumption is often increased, oxygen consumption is severely decreased after I/R injury. Based on these observations, we hypothesized that I/R injury would result in systemic organ VO2-DO2 alterations, which are distinct from those that occur in sepsis. MATERIALS AND METHODS: We used the in situ autoperfused feline ileal preparation to simultaneously examine microvascular permeability, reflected as the ileal lymph to plasma protein concentration ratio (CL/CP), and ileal VO2-DO2 relationships after either intravenous LPS (2.0 mg/kg; n = 5) or I/R injury (n = 5), and in matching controls (n = 5). RESULTS: As expected, all LPS-treated and I/R-injured animals were found to have extensive ileal histological damage and marked increases in the CL/CP compared with controls (0.315 +/- 0.009 and 0.329 +/- 0.034, respectively, v 0.097 +/- 0.009; P < .001, both comparisons). In addition, the critical DO2 (DO2c) was elevated, and the critical oxygen extraction was decreased in both the I/R and LPS groups relative to controls. However, as initially hypothesized, the VO2 at the critical DO2 was markedly decreased in the I/R group compared with that of the LPS group. CONCLUSIONS: These data indicate that I/R injury is insufficient to account for the systemic organ VO2-DO2 alterations that occur with LPS injury.     

Amino-terminal charge affects the periplasmic accumulation of recombinant heregulin/EGF hybrids exported using the Escherichia coli alkaline phosphatase signal sequence

An Escherichia coli expression system that exploits the bacterial alkaline phosphatase (PhoA) signal sequence to translocate recombinant human epidermal growth factor (hEGF) to the periplasm was used to evaluate how changes in the composition and sequence of amino acids near the PhoA-hEGF junction influence the periplasmic accumulation of recombinant protein. A series of chimeric structural genes was generated by in vitro replacement of hEGF sequence with analogous segments from the EGF-like domain of human heregulin (HRG), significantly altering the electrostatic character of the amino-terminal region of the mature protein. Quantitation of HRG/EGF protein in E. coli periplasmic extracts, by RP-HPLC, showed a fourfold decrease after one of two acidic residues located in the amino-terminal region of the mature hEGF, near the PhoA junction, was replaced. An additional threefold decrease was observed when the second acidic residue was replaced with a positively charged lysine. Further extension of the amino-terminal HRG sequence, beyond the first six residues, resulted in net neutralization of a more distant EGF acidic residue with no additional effect on protein yield. The importance of having a negatively charged group in the amino-terminal region of the mature protein was confirmed when insertion of an aspartic acid near the amino-terminus of two poorly expressed hybrid protein sequences resulted in a five- to eightfold increase in their recovery from the periplasm. This study demonstrates the importance of having negatively charged residues near the fusion junction of recombinant proteins expressed in E. coli using the PhoA signal sequence for protein export.     

Auditory behavior monitoring after bacterial meningitis. Case report

Bacterial meningitis is the main cause for acquired hearing loss. Nevertheless very little has been written about the development of the auditory behaviour either for improvement or for deterioration, after hospital release. The present study describes the case of a five month old boy with Haemophilus influenzae meningitis. Amongst various complications, a decrement in the auditory acuity was detected in the immediate evolution, with significant improvement later on by qualitative and quantitative tests.     

Metastasis induced by overexpression of p185neu-T after orthotopic injection into a prostatic epithelial cell line (NbE)

The effects of the composition of bacteriological growth media on the light output in a chemiluminometric assay of beta-galactosidase in Escherichia coli using 1,2-dioxetane substrates has been studied. In this assay a basic conflict exists between conditions that promote optimal bacterial growth and those conducive to maximal chemiluminescence. Common medium ingredients such as yeast or beef extract, protein hydrolysates and lactose suppress light emission and/or lead to high backgrounds. Quenching of light emission is probably partly due to light absorption by medium ingredients such as oxgall, and partly to interference with the reaction triggering the chemiluminescent process. Elevated backgrounds are caused by the presence of high concentrations of protein hydrolysates, which interact with the alkali in the accelerator solution. Only two purposely developed media, i.e. ILM and Colicult are shown to reconcile the requirements of growth support with that of optimal luminescent properties.     

Pharmacologic control of a humanized gene therapy system implanted into nude mice

Systemic delivery of specific therapeutic proteins by a parenteral route of administration is a recognized practice in the management of several gene defects and acquired diseases. As an alternative to repetitive parenteral administration, gene therapy may provide a novel means for systemic delivery of therapeutic proteins while improving patient compliance and therapeutic efficacy. However, for gene therapy to be an efficacious and safe approach to the clinical management of such diseases, gene expression must be tightly regulated. These investigations demonstrate precise in vivo control of protein expression from cells that are engineered to secrete human growth hormone (hGH) in response to stimulation by rapamycin. The cells were implanted intramuscularly into nu/nu mice and stimulated by intravenous or oral administration of rapamycin. In vivo experiments demonstrate that the activity and pharmacokinetics of rapamycin determine the level of serum hGH that result from the engineered cells. In addition, responsiveness of the cells to rapamycin, number of cells implanted, hGH expression kinetics, and the pharmacokinetics of hGH itself, also influence the circulating levels of hGH after rapamycin stimulation. Controlled manipulation of several of these parameters, either independently or in combination, allows for precise regulation of circulating hGH concentration in vivo.     

Calcium phosphate/calcium oxalate crystal association in urinary stones: implications for heterogeneous nucleation of calcium oxalate

PURPOSE: Most stones contain more than one type of crystals, and some combinations, such as calcium phosphate/calcium oxalate, are more common than others. Epitaxy between the crystals has been suggested to play a role in growth of such stones. The specific aim of this study is to investigate the involvement of calcium phosphate in crystallization of calcium oxalate. MATERIALS AND METHODS: Twenty calcium oxalate stones or stone fragments were examined using various microscopic techniques, including scanning, transmission and back-scattered electron microscopy. Similarly, calcium oxalate stones induced on a plastic foreign body implanted inside urinary bladders of laboratory rats were also investigated. Examination of the interface between calcium phosphate and calcium oxalate crystals was emphasized. RESULTS: Close association between crystals of calcium phosphate and calcium oxalate were found in both the human and rat stones. All crystals examined were associated with an organic matrix on the surface and contained copious amounts of organic material within the crystalline entities. Interface between the crystals also appeared to be occupied by organic matrix. CONCLUSIONS: Results of this and other studies from our laboratory indicate that epitaxy between various crystals, even though theoretically possible, appears unlikely in vivo. The appearance of specific crystalline combinations in stones is probably a result of the urinary environment being conducive for crystallization of those components. Heterogeneous nucleation of calcium oxalate is most probably induced by biological elements, including membranous cellular degradation products.