Cancer cachexia results in an increase in TNF-alpha receptor gene expression in both skeletal muscle and adipose tissue

The mRNA content of both TNF-alpha and its receptors (TNFR1 and TNFR2) in skeletal muscle and adipose tissue from cachectic rats has been assessed. The implantation of the Yoshida AH-130 ascites hepatoma resulted in substantial decrease in skeletal muscle TNF-alpha expression as early as at day 2 following tumour implantation. However, the mRNA content for the two receptors followed a different pattern, being significantly increased at day 7. In adipose tissue the expression of the TNF-alpha gene was significantly increased at all the time points studied, whereas TNF-alpha receptors expression followed a similar pattern to that observed in skeletal muscle. Western blot analysis indicated that the TNFR1 protein followed an identical pattern to that observed in the mRNA content both in muscle and adipose tissue. It is concluded that, during experimental cancer cachexia, the contribution of muscle-produced TNF-alpha is decreased; however, significant changes were observed in relation with TNF-alpha receptors at the skeletal muscle level that could possibly be related to the muscle wasting process associated with tumour growth.     

Changes in the neuromuscular synapse induced by an antibody against gangliosides

In this study, we used a monoclonal IgM antibody from a patient with a pure motor chronic demyelinating polyneuropathy, which binds specifically to the complex gangliosides GM(2), GalNAc-GD(1a), and GalNAc-GM(1b), which appear to have a common epitope of -[GalNAcbeta1-4Gal(3-2alphaNeuAc)beta1]. This was done for the following reasons: (1) to localize these gangliosides in specific cellular components of the neuromuscular junction (NMJ), and (2) to describe the anti-ganglioside antibody-induced structural and functional changes in the NMJs to gain insight into the role of gangliosides in the synaptic function. Using immunofluorescence techniques, we found that these gangliosides are located only in the presynaptic component of the motor end-plates, both in nerve terminals and in Schwann cells. After 2 weeks of continued passive transfer of the IgM monoclonal antibody over the mouse levator auris longus muscle, electromyography showed an axonal or NMJ disorder. Morphology showed important nerve terminal growth and retraction changes. Using intracellular recording electrophysiology, we found neurotransmitter release alterations, including quantal content reduction and an immature expression of voltage-dependent calcium channels similar to what occurred during NMJ development and regeneration. These changes were complement independent. The results showed that these gangliosides were involved in the reciprocal Schwann cell-nerve terminal interactions, including structural stability and neurotransmission.     

Purine level regulation during energy depletion associated with graded excitatory stimulation in brain

OBJECTIVES: The formation and release of adenosine following graded excitatory stimulation of the brain may serve important physiological functions such as sleep regulation, as well as an early resistance mechanism against excitotoxicity. However, adenosine at high levels may reflect merely the results of obstructed energy metabolism. METHODS: We examined the extent to which levels of adenosine and adenylate energy charge are affected in vivo by graded excitatory stimulations of brain using unilateral intrastriatal injections of glutamatergic agents and head-focused high energy microwaving for accurate and precise measures of purines. RESULTS: Our results confirmed that adenosine levels rise when adenylate energy charge decreases and showed that these increases occurred in three distinct phases with the rate of adenosine formation in each phase increasing as tissue adenylate energy charge was further depleted. In addition, we observed that, in most cases, the effects of focal excitatory stimulation on changes in tissue purine levels were restricted spatially within the immediate vicinity of the injection site; however, when strongly depolarizing stimuli were used, changes in purine levels could be observed in adjacent and, occasionally, even in contralateral brain regions. DISCUSSION: These results provide new insight into purine regulation that occurs under physiologically relevant conditions, such as sleep and during the early stages of brain insults that induce excitotoxicity.     

Relationship between clinical and environmental isolates of Pseudomonas aeruginosa in a hospital setting

BACKGROUND: Populations of Pseudomonas aeruginosa have been extensively studied, although there is no general agreement concerning their genetic structure. It has been proposed that P. aeruginosa is a very homogeneous species with 90% of individuals within the same clonal group; nonetheless, other results suggested that Pseudomonas populations are panmictic. Here we compared P. aeruginosa populations from clinical and environmental samples, both isolated from the Bellvitge Hospital of the University of Barcelona in Spain. METHODS: Antibiotic susceptibility determination as well as whole cell and outer membrane protein denaturing gel electrophoresis, pulsed-field electrophoresis, and random amplified polymorphic DNA analysis were performed. RESULTS: Environmental isolates were much more susceptible to antibiotics than those isolated from clinical specimens. The remainder of the analyses revealed high degree of diversity. CONCLUSIONS: Whole-cell proteins, outer-membrane proteins, and pulsed field electrophoresis did not support a close relationship between clinical and environmental isolates. Random amplified polymorphic DNA (RAPD) confirmed the distance between isolates from both sources. This suggests that the origin of hospital infections by P. aeruginosa is due mainly to growth of bacterial strains acquired by patients prior to hospital admission or from patient-to-patient through healthcare workers (HCWs).     

Plasma from pregnant rats has anti-tumoural activity

Plasma from 12-day pregnant rats showed a marked anti-tumoural activity when added to short-term cultures of rat Yoshida AH-130 cells. Indeed, 24 h after the addition of 10% pregnant rat plasma resulted in a 34% decrease in cell number as compared with cells without any serum additions or those that were added with virgin plasma. The decrease in cell number was associated with the presence of a marked aneuploid peak, as indicated by flow cytometry analysis, which suggests that pregnant plasma induces apoptosis in this particular tumour cell line. Interestingly, addition of human chorionic gonatrophin to the cell cultures did not exert any effects either on cell content of cell cycle distribution, therefore suggesting that the presence of high concentrations of this hormone in pregnant plasma is unlikely to be the cause of the increased cell death.     

Increased expression of the cyclin-dependent kinase inhibitor p27 in cleavage-stage human embryos exhibiting developmental arrest

It is accepted that approximately 50% of embryos obtained after IVF arrest during the first week. Traditionally, chromosome abnormality and suboptimal culture conditions have been proposed as factors commonly associated with embryo arrest. However, even when considering 'ideal' conditions and embryos of only excellent morphology in vitro, there is still a significant incidence of embryonic arrest. There is considerable evidence that the nuclear protein p27, a member of the Cip/Kip family of CDK inhibitors, plays an important role in multiple fundamental cellular processes, including cell proliferation, cell differentiation, and apoptosis. The present investigation, using immunocytochemical techniques coupled with confocal microscopy, was undertaken to determine whether p27 could play a role in the arrest of 4-8-cell human embryos. A total of 28 preimplantation embryos at the 4-8-cell stage were investigated. Of these, 16 were diploid embryos showing cleavage arrest with no further progression, and 12 were normally developing embryos. There was a 2-fold increased expression of the cell-cycle inhibitor p27 in arrested embryos compared with control normally developing embryos. This study represents the first demonstration of an increased expression of p27 in cleavage-stage human arrested embryos.     

Mechanism of action and resistance to monoclonal antibody therapy

Monoclonal antibodies (MoAbs) are increasingly used in the treatment of patients with hematological malignancies and autoimmune diseases. The most commonly employed humanized and chimeric MoAbs are rituximab, alemtuzumab (Campath-1H, Ilex Pharmaceuticals, San Antonio, TX), and gemtuzumab-ozogamicin (Mylotarg, Wyeth-Ayerst Laboratories, St Davids, PA). The mechanism of action of these antibodies, and host and cellular factors influencing the response, are not completely known. Induction of apoptosis, antibody-dependent cell cytotoxicity (ADCC), and complement-mediated cell death (CDC) is the proposed mechanism of action of these antibodies. We review the current understanding of the mechanism of action of and resistance to these MoAbs.     

Incidence and clinical significance of bcl-2/IgH rearrangements in follicular lymphoma

Bcl-2/IgH rearrangement is the molecular hallmark of follicular lymphoma (FL) which is present in 70-90% of the cases at diagnosis. The clinical significance of this feature is controversial. The aim of this study was to analyze the bcl-2/IgH rearrangement by means of a PCR technique, and to correlate molecular findings with clinical characteristics and outcome. Sixty-nine patients (median age, 53 years; male/female ratio: 35/34) diagnosed with FL in a single institution were included in the study. A total of 77 DNA samples were analyzed, 54 were obtained from lymph node biopsy and 23 from peripheral blood or bone marrow. Bcl-2/IgH rearrangement was assessed for both the major breakpoint region (MBR) and the minor cluster region (mcr) breakpoints by a PCR technique. Thirty-nine out of sixty patients (65%) with assessable samples were found to have a bcl-2/IgH rearrangement in the MBR breakpoint, whereas bcl-2/IgH rearrangement in mcr was observed in one patient (2%) and no rearrangement at MBR or mcr in the remaining 20 patients (33%). Regarding the initial characteristics, patients with bcl-2/IgH rearrangements at MBR or mcr were younger (<65 years) than those with no rearrangement at these sites (p = 0.0001). No differences were found according to bcl-2/IgH rearrangement in terms of complete response rate, time to treatment failure and overall survival. In our series bcl-2/IgH rearrangement at MBR or mcr, which was found in 67% of the patients, was not correlated with response to treatment, survival nor time-to-treatment-failure.