Limbic network interactions leading to hyperexcitability in a model of temporal lobe epilepsy

In mouse brain slices that contain reciprocally connected hippocampus and entorhinal cortex (EC) networks, CA3 outputs control the EC propensity to generate experimentally induced ictal-like discharges resembling electrographic seizures. Neuronal damage in limbic areas, such as CA3 and dentate hilus, occurs in patients with temporal lobe epilepsy and in animal models (e.g., pilocarpine- or kainate-treated rodents) mimicking this epileptic disorder. Hence, hippocampal damage in epileptic mice may lead to decreased CA3 output function that in turn would allow EC networks to generate ictal-like events. Here we tested this hypothesis and found that CA3-driven interictal discharges induced by 4-aminopyridine (4AP, 50 microM) in hippocampus-EC slices from mice injected with pilocarpine 13-22 days earlier have a lower frequency than in age-matched control slices. Moreover, EC-driven ictal-like discharges in pilocarpine-treated slices occur throughout the experiment (< or = 6 h) and spread to the CA1/subicular area via the temporoammonic path; in contrast, they disappear in control slices within 2 h of 4AP application and propagate via the trisynaptic hippocampal circuit. Thus, different network interactions within the hippocampus-EC loop characterize control and pilocarpine-treated slices maintained in vitro. We propose that these functional changes, which are presumably caused by seizure-induced cell damage, lead to seizures in vivo. This process is facilitated by a decreased control of EC excitability by hippocampal outputs and possibly sustained by the reverberant activity between EC and CA1/subiculum networks that are excited via the temporoammonic path.     

K-ras gene mutational analysis supports a monoclonal origin of biphasic pleomorphic carcinoma of the lung.

We investigated 27 pleomorphic carcinomas of the lung for exon 1 K-ras gene mutations using polymerase chain reaction-single-strand conformation polymophism analysis and direct sequencing. All pleomorphic carcinomas were biphasic, that is, composed of an adeno-, squamous- or large-cell-carcinomatous component associated with a spindle- and/or giant-cell component. Of 27 cases, six (22%) showed K-ras codon 12 mutations, which is a figure higher than that previously reported on in pure sarcoma-like pleomorphic carcinomas. Five tumors displayed the same mutation in both the epithelial and the sarcomatoid components, whereas in one tumor the mutation was restricted to the epithelial component. All mutations occurred in smokers, and were transversions, including GGT (glycine) to TGT (cysteine) change in two cases, to GCT (alanine) in two and to GTT (valine) in two. No significant relationships were found between the occurrence and type of mutations and patients' survival or any other clinicopathological variable, suggesting that K-ras mutations are early events in the development of these tumors. Our results indicate that most, though not all, biphasic pleomorphic carcinomas of the lung are monoclonal in origin, and that cigarette smoking may have a causative role in the development of K-ras alterations in these tumors, as all mutations are transversions.     

P53 and cathepsin-d are independent of established prognostic factors in breast-cancer

p53 and cathepsin D expression was investigated in 300 primary breast cancers by the avidin-biotin immunoperoxidase method using two murine monoclonal antibodies: PAb1801 and anti-procathepsin D, respectively. The frequency of p53- and cathepsin D-positive cells varied widely among different tumors and most tumors (82% and 86%) at least occasionally showed positive cells. The two biological markers were unrelated to one another, to cell proliferative rate and ploidy and were differently related to other biological and pathological features. In particular, p53 was directly related to tumor size and nodal involvement and inversely related to the presence of steroid receptors. Conversely, cathepsin D was directly related only to nodal involvement.     

Value of P-glycoprotein and clinicopathologic factors as the basis for new treatment strategies in high-grade osteosarcoma of the extremities.

PURPOSE: To evaluate the prognostic value of P-glycoprotein and clinicopathologic parameters in a large series of high-grade osteosarcoma (OS) patients treated at the Rizzoli Institute. PATIENTS AND METHODS: With the use of immunohistochemistry, P-glycoprotein was assessed in 149 patients with primary, nonmetastatic, high-grade OS who were homogeneously treated with chemotherapy protocols based on doxorubicin, high-dose methotrexate, and cisplatin and the addition of ifosfamide in the postoperative phase. RESULTS: P-glycoprotein positivity was found in 47 of 149 cases (32%) and was significantly associated with a higher incidence of relapse and a worse outcome, as was age younger than 12 years and tumor volume greater then 150 mL at diagnosis. Multivariate analysis further confirmed the prognostic value of these parameters, which all were independent adverse prognostic factors. Event-free survival and proportional hazards regression analyses confirmed that overexpression of P-glycoprotein at clinical onset is the most important adverse prognostic factor for high-grade OS patients treated with these chemotherapy protocols. CONCLUSION: Increased P-glycoprotein levels, together with tumor volume and age, should be taken into consideration to identify, at time of diagnosis, subgroups of OS patients with a higher risk of recurrence. This subgroup identification will constitute the basis for drawing individualized treatment protocols on the basis of risk evaluation, with the aim of using more aggressive chemotherapy, or combination chemotherapy with other adjuvants, only in those patients for which more aggressive regimens are strictly necessary and warranted.     

The intestinal ecosystem in chronic functional constipation

Chronic functional constipation is common in infants, and the bacterial composition of stools in this condition is not known. The study aims were to: (i) investigate the composition of the intestinal ecosystem in chronic functional constipation; (ii) establish whether the addition of the water-holding agent calcium polycarbophil to the diet induces an improvement in constipation; and (iii) determine the composition of the intestinal ecosystem after the use of this agent. In total, 42 children (20F, 22M; mean age: 8.6±2.9y) were studied. Twenty-eight children with functional chronic constipation without anatomical disorders were treated double-blind in random sequence for 1 month with an oral preparation of calcium polycarbophil (0.62g /twice daily) or placebo. Intestinal flora composition was evaluated by standard microbiological methods and biochemical assays on faecal samples collected before and after treatment. Fourteen healthy children were studied as controls. The results show that (i) the constipated children presented a significant increase in clostridia and bifidobacteria in faeces compared to healthy subjects-different species of clostridia and enterobacteriaceae were frequently isolated; no generalized overgrowth was observed; Clostridia outnumbered bacteroides and E. coli mean counts by 2-3log, while bacteroides and E. coli counts were similar (5-6 log10/g fresh faeces); these intestinal disturbances could be defined as a dysbiosis, i.e. a quantitative alteration in the relative proportions of certain intestinal bacterial species. (ii) Clinical resolution of constipation was achieved only in 43% of treated children and an improvement in 21% (one bowel movement every 2 d). (iii) Calcium polycarbophil treatment induced no significant changes in the composition of the intestinal ecosystem, nor in blood chemistry parameters.     

Effects on Serum Inflammatory Cytokines of Cholecalciferol Supplementation in Healthy Subjects with Vitamin D Deficiency

The effects of different cholecalciferol supplementation regimens on serum inflammatory cytokines in healthy subjects with vitamin D deficiency are still lacking. This is a single-center, open-label, randomized, parallel group study involving healthy subjects deficient in vitamin D (baseline 25OHD < 20 ng/mL) receiving oral cholecalciferol with three different dosing regimens: Group A: 10,000 IU/day for 8 weeks followed by 1000 IU/day for 4 weeks; Group B: 50,000 IU/week for 12 weeks and Group C: 100,000 IU every other week for 12 weeks. IL-17A, IL-6, IL-8, IL-10, IL-23 and TNFα were measured at baseline and at week 4, 8, 12, and 16. 75 healthy subjects were enrolled (58.7% female), with an average age of 34.1 ± 10.2 years. No statistical differences were observed among groups at baseline for either IL-6, IL-17A, IL-23, IL-8 or IL-10 at any time point; TNFα was indetectable. Concerning the whole sample, the time trend analysis showed a statistically significant linear trend for decreasing values over the treatment period for IL-6 (p = 0.016) and IL-17A (p = 0.006), while no significant time trends were observed for the other teste cytokines. No significant differences were found in the serum concentrations of the tested cytokines between week 12 and week 16. In young healthy individuals deficient in vitamin D, cholecalciferol administration showed a decrease in the serum IL-6 and IL-17A concentrations, without marked differences using the three regimens.