Stem cell marker nestin and c-Jun NH2-terminal kinases in tumor and peritumor areas of glioblastoma multiforme: possible prognostic implications.

PURPOSE: It has been hypothesized that brain tumors are derived from stem cell or transiently dividing precursor transformation. Furthermore, c-Jun NH(2)-terminal kinases (JNKs) have been involved in gliomagenesis. This study analyzes stem cell marker nestin and JNK expression in glioblastoma multiforme (GBM) and peritumor tissue and assesses their possible prognostic implications. EXPERIMENTAL DESIGN: Nestin and both total JNK (tJNK) and phosphorylated JNK (pJNK) expression was investigated by immunohistochemistry in 20 GBMs. Samples were derived from tumors (first area), from tissues at a distance <1 cm (second area), and between 1 and 3.5 cm (third area) from the macroscopic tumor border. The relationships between patients' age, Karnofsky performance status, gender, protein expression, and survival were analyzed. RESULTS: Nestin cytoplasmic immunoreactivity was observed in the majority of cells in tumor but infrequently in peritumor areas. tJNK, observed in the nucleus and cytoplasm, was widely expressed in the three areas; pJNK, mostly located in the nuclei, was found in a variable percentage of cells in the tumor and peritumor tissue. Nestin and JNK expression in peritumor areas was independent of the presence of neoplastic cells. Univariate analysis indicated that survival was longer (19 versus 12 months; P = 0.01) for patients whose pJNK/nestin and (pJNK/tJNK)/nestin ratios in the second area were > or =2.619 and > or =0.026, respectively. The same variables showed an independent prognostic value in multivariate analysis. CONCLUSIONS: Nestin and JNK expression indicates that peritumor tissue, independently of the presence of neoplastic cells, may present signs of transformation. Moreover, pJNK/nestin and (pJNK/tJNK)/nestin ratios in that tissue seem to have some prognostic implications in GBM patients.     

Goserelin as ovarian protection in the adjuvant treatment of premenopausal breast cancer: a phase II pilot study

The aim of the present trial was to investigate the protective effects on ovarian function, and the efficacy and tolerability of goserelin added to adjuvant chemotherapy for early breast cancer. Following surgical treatment, 64 premenopausal patients with early breast cancer received goserelin 3.6 mg (every 28 days for 1 year) and an adjuvant treatment which was chosen according to the patient's prognosis. Median age was 42 years (range 27-50). ECOG performance status was 0-1 in all patients. Twenty-eight patients (44%) had estrogen receptor (ER)+ tumors and 36 (56%) patients had ER- tumors. Fifty-two (81%) patients had stage II disease and 12 (19%) had stage III disease. Eighteen patients received cyclophosphamide, methotrexate and 5-fluorouracil chemotherapy, 46 patients received an anthracycline-based regimen, and nine of them received high-dose chemotherapy and autologous peripheral blood progenitor cell transplantation. Fifty-one patients (80%) were irradiated. ER+ patients also received tamoxifen for 5 years. Serum estradiol was suppressed to values below 40 pg/ml in all patients. After a median follow-up of 55 months, 86% of patients had resumed normal menses, 84% of patients were disease-free and 94% were alive. The 1-, 3- and 5-year projected recurrence-free survival rates were 100, 81 and 75%, respectively. Five years after treatment one patient had a pregnancy that ended with a normal childbirth. No unexpected adverse events were reported. These data show that the addition of goserelin to adjuvant therapy of premenopausal patients with early breast cancer is well tolerated and protects long-term ovarian function.     

Fos induction in cortical interneurons during spontaneous wakefulness of rats in a familiar or enriched environment

It has been repeatedly reported that Fos is spontaneously induced in several brain structures, including the cerebral cortex, during wakefulness. To ascertain whether cortical interneurons are involved in this state-dependent oscillation of gene regulation, we combined Fos immunocytochemistry with immunostaining of either parvalbumin or calbindin, known markers of cortical interneurons. Immunopositive neurons were examined in the sensorimotor and cingulate cortex. In rats perfused in basal conditions, a minor proportion (around 8%) of Fos-immunoreactive neurons in the parietal cortex were also parvalbumin- or calbindin-immunoreactive; these double immunostained cells accounted for 13% of the parvalbumin- and 34% of the calbindin-labeled neurons. Colocalization of Fos with either calcium-binding protein was instead not observed in the cingulate cortex. In rats stimulated by novel environmental cues during the period of wakefulness preceding perfusion, Fos-positive neurons increased markedly relative to unstimulated animals, and involved the majority of the calbindin- or parvalbumin-labeled cell populations (60-75% and over 95%, respectively). In the neuronal populations in which Fos was induced by exposure to the enriched environment, the proportion of calbindin- and parvalbumin-labeled cells was larger than in the unstimulated cases, and the increment was statistically significant in the cingulate cortex. The results demonstrate that Fos induction occurring in the cortex during undisturbed wakefulness in a familiar environment involves a minor proportion of interneurons. Furthermore, the findings indicate that the addition of novel environmental stimuli results in an increase of Fos-expressing neurons whose recruitment, at least in the cingulate cortex, involves a higher proportion of interneurons than of projection neurons.     

Microglia activation and cell death in response to diethyl-dithiocarbamate acute administration

An increasing body of evidence suggests a role for activated microglia in the pathogenesis of neurodegenerative disorders. Hence, it would be useful to have a better understanding of the significance of microglial activation for neuronal damage. Unfortunately, most models of microglial activation use invasive or long-lasting insults, which make it difficult to evaluate the role played by microglia. We have instead developed a model for microglial activation by using brief exposure to the widely available neurotoxin diethyl-dithiocarbamate (DDTC). Despite evidence for the neurotoxic nature of this substance, microglia involvement has not been hitherto investigated. After acute i.p. administration of DDTC at two different doses, microglia were already activated in selected areas of the rat brain (hippocampal dentate gyrus, entorhinal-pyriform cortex and hypothalamus) after 1 hour, reaching a peak at 3-6 hours and subsided within 6-48 hours, depending on the brain region. Microglia activation was associated with interleukin-1 beta immunopositivity between 3 and 6 hours and with up-regulation of major histocompatibility complex class II expression between 24 and 48 hours. No significant changes in astrocyte immunostaining were detected between 6 hours and 6 days. The TUNEL procedure revealed the death of a limited number of cells in the above-mentioned structures that peaked at 6h and then declined rapidly. Cell death was detected in sites with major, minor, or no microglial activation, indicating that these two events can occur concomitantly or independently. The study shows that the administration of DDTC provides a useful model for studying the implications of region-specific reactivity of microglia and its differential interaction with neuronal damage.     

Lack of apoptosis in patients with progressive external ophthalmoplegia and mutated adenine nucleotide translocator-1 gene

Adenine nucleotide translocator-1 (ANT-1), encoded by chromosome 4 (4q34-35 locus), is a component of the mitochondrial permeability transition pores that are involved in apoptotic mechanisms. We studied muscle biopsies from seven individuals with autosomal dominant progressive external ophthalmoplegia caused by ANT-1 mutations. We found no instance of terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling (TUNEL) positivity nor significant expression of apoptosis-related proteins. Furthermore, there was no morphological evidence of apoptosis at the ultrastructural level. Thus, degeneration of muscle in this disorder is nonapoptotic.     

A case of CPT deficiency, homoplasmic mtDNA mutation and ragged red fibers at muscle biopsy

A 45-year-old male patient had an episode of acute renal failure with myoglobinuria, myalgias, weakness, and markedly increased serum CK levels. Similar episodes had occurred in the past. Carnitine palmitoyl-transferase II (CPT II) deficiency was documented both biochemically and genetically. Interestingly, muscle biopsy also showed some ragged red fibers (RRF) and complete mitochondrial DNA (mtDNA) sequence disclosed a homoplasmic T3394C point mutation. This mutation is described in Leber's hereditary optic neuropathy (LHON) or in patients with diabetes mellitus.     

Immunomodulating effect of antimicrobial agents on cytokine production by human polymorphonuclear neutrophils

It has been previously demonstrated that some antimicrobial agents enhance activities of human polymorphonuclear neutrophils (PMNs). The effect on the release of cytokines in an inflammatory context from PMNs by these antibiotics was evaluated. We studied the effect of the release of some cytokines by human PMNs RT-PCR analysis on a clinical strain of Klebsiella pneumoniae by comparing the effect with that observed in the presence of co-amoxiclav, sanfetrinem, clarithromycin, prulifloxacin and tobramycin. All the drugs tested were capable of modulating PMN synthesis in vitro of pro-inflammatory cytokines IL-8, IL-1beta, TNF-alpha and IL-6, but not that of anti-inflammatory cytokine IL-10. The degree of their stimulatory or inhibitory potency varied with the cytokine examined.     

Acetylcholinesterase induces the expression of the beta-amyloid precursor protein in glia and activates glial cells in culture

Acetylcholinesterase (AChE) activities in CNS physiopathology are increasingly diverse and range from neuritogenesis, through synaptogenesis, to enhancement of amyloid fiber assembly. In Alzheimer's disease, senile plaques and neurodegeneration specially affect regions enriched for cholinergic synapses. In this study we show an effect of AChE that could contribute to the increased deposition of Abeta in certain regions. Affinity-purified AChE induced the expression of amyloid-beta-precursor protein (beta-APP) in glial cells in a concentration-dependent manner up to 5 nM. In glia, AChE also increased inducible nitric oxide synthase (iNOS) assessed by immunocytochemistry and decreased reductive metabolism as evidence of cell activation. AChE could increase the expression of beta-APP in astrocytes and microglia as result of the activation of glial cells. As a whole, we found that AChE has additional effects that could result in an increased synthesis of Abeta, both by increasing beta-APP expression of astrocytes and by further activating glial cells.