Comparison of Two Culture Methods for Detection of Tobramycin-Resistant Gram-Negative Organisms in the Sputum of Patients with Cystic Fibrosis

A culture method utilizing quantitative plating on antibiotic-containing media has been proposed as a technique for the detection of tobramycin-resistant organisms that is more sensitive than standard methods. Typical sputum culture methods quantitate the relative amounts of each distinct morphotype, followed by antibiotic susceptibility testing of a single colony of each morphotype. Sputum specimens from 240 cystic fibrosis patients were homogenized, serially diluted, and processed in parallel by the standard method (MacConkey agar and OF basal medium with agar, polymyxin, bacitracin, and lactose) and by plating on antibiotic-containing media (MacConkey agar with tobramycin added at 25 microg/ml [MAC-25] and 100 microg/ml [MAC-100]). MICs of tobramycin were determined for all Pseudomonas aeruginosa isolates by broth microdilution. Growth of P. aeruginosa on MAC-25 was considered to be equivalent to a tobramycin MIC of > or = 16 microg/ml, and growth on MAC-100 was considered to be equivalent to a tobramycin MIC of > or = 128 microg/ml. Analysis of method-specific detection rates showed that tobramycin-containing medium was more sensitive than the standard method for the detection of tobramycin-resistant P. aeruginosa, Stenotrophomonas maltophilia, and Achromobacter xylosoxidans but was less sensitive for the detection of Burkholderia cepacia than the standard method. When MICs for P. aeruginosa that grew on tobramycin-containing medium were tested by broth microdilution, the MICs for 28 of 121 strains (23%) growing on MAC-25 and 22 of 56 strains (39%) growing on MAC-100 were MICs < 16 and < 128 microg/ml, respectively. Addition of a tobramycin-containing MacConkey plate to the routine media for sputum culture may provide additional, clinically relevant microbiologic data.     

Chromosomal integrity maintained in five human embryonic stem cell lines after prolonged in vitro culture

There have been recent reports of human embryonic stem cell (hESC) lines developing chromosomal aberrations after long-term culture, indicating an unstable genomic status due to the in vitro milieu. This raises concern, since it would limit their use in therapeutics. In this study the chromosomal status of five well-characterized hESC lines, SA002, SA002.5, AS034.1.1, SA121 and SA461, was monitored during long-term in vitro culture. The criteria of defined hESCs were met by all of the five hESC lines (four diploid and one trisomic for chromosome 13). The genomes were screened for chromosomal aberrations and rearrangements using comparative genomic hybridization (CGH), interphase fluorescence in situ hybridization (FISH) and traditional karyotyping on several occasions while in culture. The genomic integrity was shown to be maintained after repeated freeze-thaw procedures and continuous culture in vitro for up to 22 months (148 passages). We discuss the most common de novo chromosomal aberrations reported in hESCs, as well as their possible origin.     

High expression of MDR1, MRP1, and MRP3 in the hepatic progenitor cell compartment and hepatocytes in severe human liver disease

An increase in bile ductular structures is observed in diverse human liver diseases. These structures harbour the progenitor cell compartment of the liver. Since ATP-binding cassette (ABC) transporters may have a cytoprotective role in liver disease, an immunohistochemical study was performed on human liver specimens from patients with primary biliary cirrhosis (PBC), chronic hepatitis C virus (HCV) infection, submassive cell necrosis, and normal liver. The expression of MDR1, MDR3, BSEP, MRP1, MRP2, and MRP3 was determined using specific antibodies. Dilution series were constructed to determine the critical staining level in order to estimate the factor of up-regulation. In normal liver, hepatocytes showed canalicular staining for MDR3, BSEP, and MRP2. MDR1 stained the canalicular membrane of hepatocytes as well as that of cholangiocytes. MRP3 showed low immunoreactivity of bile duct epithelial cells and centrilobular hepatocytes only. Normal liver showed no immunoreactivity for MRP1. In diseased liver, the expression of MDR3, BSEP, and MRP2 was relatively stable. In PBC, HCV, and submassive necrosis, the expression levels of MDR1, MRP1, and MRP3 were increased. The strongest immunoreactivity was seen after submassive necrosis, where remaining islands of hepatocytes showed strong canalicular staining for MDR1 and MRP3. Regenerating bile ductules at the interface of portal tracts and necrotic areas stained intensely for MDR1, MRP1, and MRP3. In conclusion, MDR1, MRP1, and MRP3 are up-regulated in hepatocytes in severe human liver disease. Strong MDR1, MRP1, and MRP3 reactivity is seen in regenerating human bile ductules.     

Anxiety Profiles of Mice Selectively Bred for Intermale Aggression

In the present study we compared genetically selected aggressive (TA) and nonaggressive (TNA) male mice, as well as males from an unselected control line (SW), in three tests of anxiety: the elevated plus-maze, the light-dark box, and the staircase test. Males were tested repeatedly for 3 days. In all three tests TA males were found to be more active and less anxious than TNA males, with SW males showing more or less intermediate scores. Furthermore, repeated plus-maze testing induced anxiety-like behavior in both TA and TNA mice, whereas repeated testing in the light-dark and staircase paradigms resulted in different responses. Whereas TNA animals were found to be stable, TA animals showed habituation on the second and third day of testing. The obtained data support the conclusion that a higher-level offensive aggression entails lower level of anxiety.     

Hemagglutination by Staphylococcus aureus strains responsible for human bacteremia or bovine mastitis

Although hemagglutination by Staphylococcus aureus has been associated with the pathogenesis of bovine mastitis, this trait has not been characterized with regard to human disease. In this study, the prevalence of hemagglutination in 100 strains of S. aureus responsible for bovine mastitis or human bacteremia, was characterized. Under optimum conditions hemagglutination was noted in 23% of the bovine strains, but only 13% of human strains, leading us to conclude that this trait is not a significant virulence determinant in human systemic infection. Additional studies indicate the hemagglutinin of S. aureus strains responsible for human bacteremia is proteinaceous in character.     

HIV accessory proteins and surviving the host cell

Human immunodeficiency virus generates the accessory proteins Nef, viral infectivity factor (Vif), viral protein R, and viral protein U or viral protein X during viral replication in host cells. Although the significance of these accessory proteins is often lost in vitro, they are essential for viral pathogenesis in vivo. Therefore, these proteins have much potential as antiviral targets. Recent data reveal Vif perturbs an ill-defined antiviral pathway in host cells allowing HIV replication. These data highlight a common feature among HIV accessory proteins in manipulating the host to aid viral pathogenesis. Therefore, these new insights into Vif and other HIV accessory proteins are reviewed, emphasizing host cell interactions and new targets for therapeutic intervention.     

Changes in bone turnover during tibolone treatment

OBJECTIVES: An open study was carried out to evaluate changes in bone remodeling markers such as N-telopeptide (NTx), tartrate-resistant acid phosphatase (TRAP), total alkaline phosphatase (TAP), and bone alkaline phosphatase (BAP) during a 1-year continuous tibolone treatment in postmenopausal women. MATERIAL AND METHODS: Thirty-six postmenopausal women were recruited for receiving tibolone 2.5 mg per day for 1 year. Densitometry and determination of biochemical markers of bone metabolism in serum and urine were performed at 1, 3, 6, and 12 months. RESULTS: Comparing baseline with 12 month's values, BAP and all resorption markers decreased significantly. NTx began to decrease since the initiation of the treatment (baseline: 74.4 +/- 5.3; 1 month: 57.5 +/- 4.2; 12 months: 36.6 +/- 2.8). BAP increased at the first month (baseline: 37.3 +/- 2.1; 1 month: 42.6 +/- 3.0) but diminished in the following months (12 months: 23.1 +/- 1.5). TAP started to decrease significantly only after 6 months of treatment (baseline: 37.3 +/- 2.1; 12 months: 31.4 +/- 2.3) and TRAP after 3 months (baseline: 9.8 +/- 0.4; 6 months: 9.1 +/- 0.5; 12 months: 8.2 +/- 0.4). Normal bone mineral density at distal and ultradistal forearm was maintained during the 1-year treatment (baseline: 0.42 +/- 0.01; 12 months: 0.42 +/- 0.01 and baseline: 0.33 +/- 0.01; 12 months: 0.33 +/- 0.01, respectively). CONCLUSION: The use of tibolone 2.5 mg per day diminished progressively and significantly bone resorption and formation markers during 1-year treatment period.     

CD4 lymphocytes in the blood of HIV(+) individuals migrate rapidly to lymph nodes and bone marrow: support for homing theory of CD4 cell depletion

Recent studies have provided evidence that macrophages from Th1-prone mouse strains respond with an M1 profile, and macrophages from Th2-prone mouse strains respond with an M2 profile, characterized by the dominant production of NO or TGF-beta 1, respectively. We have shown that peritoneal macrophages from IL-12p40 gene knockout mice have a bias toward the M2 profile, spontaneously secreting large amounts of TGF-beta 1 and responding to rIFN-gamma with weak NO production. Moreover, IL-12p40KO macrophages are more permissive to Trypanosoma cruzi replication than their wild-type littermate cells. Prolonged incubation with rIL-12 fails to reverse the M2 polarization of IL-12p40KO macrophages. However, TGF-beta 1 is directly implicated in sustaining the M2 profile because its inhibition increases NO release from IL-12p40KO macrophages. IFN-gamma deficiency is apparently not the reason for TGF-beta 1 up-regulation, because rIFN-gamma KO macrophages produce normal amounts of this cytokine. These findings raise the possibility that IL-12 has a central role in driving macrophage polarization, regulating their intrinsic ability to respond against intracellular parasites.     

Mapping of gene loci in the Q13–Q15 region of chromosome 12

The gene loci CDK4, GLI, CHOP and MDM2 have been mapped to the q13–q15 region of chromosome 12. Using fluorescencein situ hybridization onto simultaneously DAPI-banded metaphase chromosomes and interphase nuclei, we have more precisely mapped and ordered these loci, together with a number of Genethon microsatellite markers. GLI and CHOP localize to 12q13.3–14.1, CDK4 to 12q14 and MDM2 to 12q14.3–q15, and the gene order is cen-GLI/CHOP-CDK4-MDM2. The Genethon microsatellites D12S80 and D12S83 flank MDM2.