Glucose may induce cell death through a free radical-mediated mechanism

It has been reported that glucose may autooxidize generating free radicals which have been hypothesized to induce important cellular abnormalities. To investigate the cell damage induced by glucose-dependent oxidative stress, the FRTL5 cell strain was incubated in 10 or 20 mM glucose, either alone or in the presence of buthionine-sulfoximine, a transition state inhibitor that blocks glutathione synthesis. We found indeed that buthionine-sulfoximine greatly inhibited glutathione production and increased malondialdehyde (a marker of oxidative cell damage) levels, especially in 20mM glucose. We also found that, when glutathione production was inhibited, 10mM glucose induced apoptosis and 20 mM glucose induced necrosis. These data show that the glucose-dependent cell damage is a function of glutathione production. They also show that such glucose-dependent free radical production may be critical for determining cell damage, even for small variations as the ones we tested (from 10 to 20 mM glucose).     

Inhibition of mitogen-activated protein kinase activity and proliferation of an early osteoblast cell line (MBA 15.4) by dexamethasone: role of protein phosphatases

Chronic glucocorticoid therapy causes rapid bone loss and clinical osteoporosis. We found that although the glucocorticoid, dexamethasone, stimulated osteoblast maturation, it also inhibited proliferation of a preosteoblastic cell line, MBA-15.4. The dexamethasone-induced decline in preosteoblast proliferation correlated with a 30-40% reduction in protein kinase C/TPA-stimulated mitogen-activated protein kinase (MAPK) activity. These steroid effects only became evident after 6-24 h treatment, implying that dexamethasone acts on de novo synthesis of proteins. Because MAPK is inactivated by dephosphorylation of tyrosine and threonine residues, cells were treated concomitantly for 24 h with dexamethasone and inhibitors of tyrosine (sodium orthovanadate) and/or serine/threonine phosphatases (sodium fluoride). MAPK activity and cell proliferation were restored when MBA-15.4 cells were treated with vanadate, suggesting that dexamethasone up-regulates tyrosine phosphatase activity. Inactivation of serine/threonine phosphatases with sodium fluoride had no effect. Inhibition of the PKA pathway (which is growth inhibitory in mature osteoblasts) with H-89 did not reverse the effects of dexamethasone. Pretreatment with dexamethasone inhibited both peak- and extended activation plateau-phases of MAPK activity. Both phases were fully restored by pretreatment with vanadate, implicating more than one tyrosine phosphatase. Cycloheximide, alone or in combination with dexamethasone, prevented drop-off from plateau to basal levels, suggesting that an inducible dual-specificity phosphatase regulates the plateau-phase. We conclude that dexamethasone may inhibit preosteoblast growth via a novel tyrosine phosphatase pathway.     

Epidermal growth factor receptors in human corpora lutea during the menstrual cycle and pregnancy

We have demonstrated the presence of epidermal growth factor (EGF) and its receptors in human non-gestational corpora lutea. To determine further the characteristics of EGF receptor binding, we examined 30 human corpora lutea throughout the luteal phase and during pregnancy. Scatchard plots of EGF binding in 29 of the 30 corpora lutea were curvilinear, suggesting negative co-operativity. The mean +/- SE of the association constant Ka was (0.9 +/- 0.2) x 10(9) l/mol, the dissociation constant Kd was (2.2 +/- 0.3) x 10(-9) mol/l and the number of binding sites (Rt) was (15.8 +/- 2.1) x 10(-19) mol/micrograms protein for non-gestational corpora lutea. The Kd increased significantly in late pregnancies compared to early pregnancies (P = < 0.005), while Rt was significantly higher in term pregnancies than in either early pregnancy (P < 0.01) or the menstrual cycle (P < 0.001). Corpora lutea atretica (n = 2) and ovarian stroma (n = 6) did not show any EGF binding activity. Our findings demonstrate the presence of specific EGF receptors in human corpora lutea of both the menstrual cycle and pregnancy. The changes in EGF binding parameters in early pregnancy suggest that there may be a relationship between the role of EGF and ovarian steroidogenesis.     

Cytokines and perinatal brain injury

A rapidly expanding body of data provides support for the hypothesis that pro-inflammatory cytokines including interleukin-1 beta (IL-1 beta), and tumor necrosis factor-alpha (TNF-alpha) are expressed acutely in injured brain and contribute to progressive neuronal damage. Little is known about the pathogenetic role of these cytokines in perinatal brain injury. Recent experimental studies have incorporated two closely related in vivo perinatal rodent brain injury models to evaluate the role(s) of pro-inflammatory cytokines in the progression of neuronal injury: a perinatal stroke model, elicited by unilateral carotid artery ligation and subsequent timed exposure to 8% oxygen in 7-day-old rats, and a model of excitotoxic injury, elicited by stereotactic intra-cerebral injection of the selective excitatory amino acid agonist NMDA. Each of these lesioning methods results in reproducible, quantifiable focal forebrain injury at this developmental stage. Acute brain injury, evoked by cerebral hypoxia-ischemia or excitotoxin lesioning, results in transient marked increases in expression of IL-1 beta, and TNF-alpha mRNA in brain regions susceptible to irreversible injury, and there is evidence that pharmacological antagonism of IL-1 receptors can attenuate injury in both models. Recent studies also suggest that complementary strategies, based on pharmacological antagonism of platelet activating factor and on neutrophil depletion can also limit the extent of irreversible injury. In summary, current data suggest that pro-inflammatory cytokines contribute to the progression of perinatal brain injury, and that these mediators are important targets for neuroprotective interventions in the acute post-injury period.     

Agreement between face-to-face and telephone-administered mood ratings in patients with rapid cycling bipolar disorder

BACKGROUND AND PURPOSE: Clinical trials have demonstrated that high dose radiation therapy and daily cisplatin (CDDP) could increase local control and survival in carcinoma from various sites. The present phase I-II study has combined high dose radiation therapy and daily CDDP at escalating dosages. METHODS: From August 1994 to December 1995, 23 patients with non-resectable carcinoma of the pancreas were enrolled in a phase I-II multicentric, pilot study to test the toxicity and the effectiveness of high dose radiotherapy and daily cisplatin (CDDP) at escalating dosages. A dose of 6 mg/sqm/day of CDDP was selected for the phase II step since no grade IV toxicity occurred in any patient in the phase I step. RESULTS: Toxicity was considered fairly acceptable. At the time of analysis, the 23 patients who entered the study had clear evidence of evolutive disease either locally or distantly in the liver. It is suggested that high dose radiotherapy (60 Gy continuously) and daily CDDP have little effect on local control of the tumor and survival, and only a moderate effect on pain. CONCLUSIONS: In unresectable, apparently non-metastatic cancers of the pancreas, there is an urgent need for new agents or new combinations of agents to be tested.     

Tissue equivalent vessel phantoms for intravascular ultrasound

The presence of the non-selective protein kinase C (PKC) inhibitors, staurosporine (100 nM) and polymyxin B (100 microM) in cultured human RPE cells for more than 24 h triggers apoptotic death. Apoptosis is characterized by a diminishing number of cells, a labelling of nuclei by the TUNEL method and by observable morphological changes. An inhibitor of PKC and cyclic nucleotide-dependent protein kinases, 1-(5-isoquinolinesulphonyl)-2-methyl piperazine (H-7; 100 microM), was without effect, as was the specific PKC inhibitor, calphostin C (100 nM). The PKC-activating phorbol esters, phorbol-12-myristate-13-acetate (PMA; 1 microM) and phorbol-12,13-dibutyrate (PDB; 1 microM) and the non-tumour-promoting phorbol ester, 4 alpha-PMA (1 microM) were without effect, as was the diacyl glycerol analogue, 1,2-dioctanoyl-snglycerol (DOG; 10 microM). The PKC activators did not attenuate the apoptosis induced by staurosporine or polymyxin B. Furthermore, deprivation of glucose and oxygen (simulated ischemia) for 72 h induced apoptosis: this could be prevented by inclusion of 10% (v/v) foetal bovine serum (FBS) but not by a variety of PKC activators. Six PKC isoenzymes were shown to be present in RPE cells (alpha, beta 1, beta 2, delta, epsilon, E) and only the calcium-dependent cPKC levels changed after treatment with staurosporine or simulated ischaemia. Since only the less selective inhibitors of PKC induced apoptosis, it is suggested that PKC is not involved directly in the induction process of apoptosis in RPE cells. It is possible that the staurosporine and polymyxin B-induced effects of apoptosis in RPE cells are triggered by an unknown kinase-dependent pathway, but whether the 'ischaemia'-induced death is related to this same process remains to be elucidated.