NOTCH1 mutations are associated with high CD49d expression in chronic lymphocytic leukemia: link between the NOTCH1 and the NF-κB pathways.

In chronic lymphocytic leukemia (CLL), stabilizing mutations of NOTCH1, affecting up to 10-15% of cases, have been associated to poor prognosis, disease progression and refractoriness to chemotherapy. NOTCH1 mutations are significantly overrepresented in trisomy 12 CLL, a disease subset frequently expressing CD49d, the α4 chain of the very-late-activation-4 integrin, a well-known key regulator of microenviromental interactions, and negative prognosticator in CLL. In the present study, by analysing a wide cohort of 1180 CLL, we observed a very strong association between the presence of NOTCH1 mutations and the expression of CD49d (P<0.0001), occurring also outside the trisomy 12 CLL subset. Using both the MEC-1 CLL-like cells stably transfected with the NOTCH1 intracellular domain and primary CLL cells bearing a mutated or wild-type NOTCH1 gene configuration, we provide evidence that triggering of the NOTCH1 pathway resulted in a positive CD49d expression regulation, which was driven by a NOTCH1-dependent activation of nuclear factot-κB (NF-κB). Consistently, pharmacological inhibition of the NOTCH1 and/or of the NF-κB pathways resulted in impaired NF-κB nuclear translocation with consequent down-modulation of CD49d expression. Altogether, our data link for the first time NOTCH1 mutations to CD49d expression regulation through the involvement of the NF-κB pathway in CLL.

Childhood neurofibromatosis type 2 (NF2) and related disorders: from bench to bedside and biologically targeted therapies.

Neurofibromatosis type 2 [NF2; MIM # 101000] is an autosomal dominant disorder characterised by the occurrence of vestibular schwannomas (VSs), schwannomas of other cranial, spinal and cutaneous nerves, cranial and spinal meningiomas and/or other central nervous system (CNS) tumours (e.g., ependymomas, astrocytomas). Additional features include early onset cataracts, optic nerve sheath meningiomas, retinal hamartomas, dermal schwannomas (i.e., NF2-plaques), and (few) café-au-lait spots. Clinically, NF2 children fall into two main groups: (1) congenital NF2 - with bilateral VSs detected as early as the first days to months of life, which can be stable/asymptomatic for one-two decades and suddenly progress; and (2) severe pre-pubertal (Wishart type) NF2- with multiple (and rapidly progressive) CNS tumours other-than-VS, which usually present first, years before VSs [vs. the classical adult (Gardner type) NF2, with bilateral VSs presenting in young adulthood, sometimes as the only disease feature]. Some individuals can develop unilateral VS associated with ipsilateral meningiomas or multiple schwannomas localised to one part of the peripheral nervous system [i.e., mosaic NF2] or multiple non-VS, non-intradermal cranial, spinal and peripheral schwannomas (histologically proven) [schwannomatosis]. NF2 is caused by mutations in the NF2 gene at chromosome 22q12.1, which encodes for a protein called merlin or schwannomin, most similar to the exrin-readixin-moesin (ERM) proteins; mosaicNF2 is due to mosaic phenomena for the NF2 gene, whilst schwannomatosis is caused by coupled germ-line and mosaic mutations either in the SMARCB1 gene [SWNTS1; MIM # 162091] or the LZTR1 gene [SWNTS2; MIM # 615670] both falling within the 22q region and the NF2 gene. Data driven from in vitro and animal studies on the merlin pathway [e.g., post-translational and upstream/downstream regulation] allowed biologically targeted treatment strategies [e.g., Lapatinib, Erlotinib, Bevacizumab] aimed to multiple tumour shrinkage and/or regression and tumour arrest of progression with functional improvement.

CD49d prevails over the novel recurrent mutations as independent prognosticator of overall survival in chronic lymphocytic leukemia.

CD49d, the alpha-chain of the integrin heterodimer α4ß1, was identified among the strongest predictors of overall survival (OS) in chronic lymphocytic leukemia (CLL), along with IGHV mutational status and deletion of the 17p chromosome involving TP53. In addition to TP53, the clinical relevance of NOTCH1, SF3B1 and BIRC3 gene mutations has been recently emphasized. By analyzing a cohort of 778 unselected CLL patients, we assessed the clinical relevance of CD49d as an OS predictor in subgroups defined by mutation/deletion of the TP53, NOTCH1, SF3B1 and BIRC3 genes. In this context, CD49d emerged as an independent predictor of OS in multivariate Cox analysis (Hazard ratio =1.88, P<0.0001). Consistently, high CD49d expression identified CLL subsets with inferior OS in the context of each category of a previously reported hierarchical risk stratification model. Moreover, by evaluating the relative importance of biological prognosticators by random survival forests, CD49d was selected among the top-ranked OS predictor (variable importance =0.0410), along with IGHV mutational status and TP53 abnormalities. These results confirmed CD49d as an independent negative OS prognosticator in CLL also in comprehensive models comprising the novel recurrent mutations. In this context, TP53 disruption and NOTCH1 mutations retained prognostic relevance, in keeping with their roles in CLL cell immuno-chemoresistance.

NOTCH1 mutations associate with low CD20 level in chronic lymphocytic leukemia: evidence for a NOTCH1 mutation-driven epigenetic dysregulation.

In chronic lymphocytic leukemia (CLL), NOTCH1 mutations have been associated with clinical resistance to the anti-CD20 rituximab, although the mechanisms behind this peculiar behavior remain to be clarified. In a wide CLL series (n=692), we demonstrated that CLL cells from NOTCH1-mutated cases (87/692) were characterized by lower CD20 expression and lower relative lysis induced by anti-CD20 exposure in vitro. Consistently, CD20 expression by CLL cells was upregulated in vitro by γ-secretase inhibitors or NOTCH1-specific small interfering RNA and the stable transfection of a mutated (c.7541-7542delCT) NOTCH1 intracellular domain (NICD-mut) into CLL-like cells resulted in a strong downregulation of both CD20 protein and transcript. By using these NICD-mut transfectants, we investigated protein interactions of RBPJ, a transcription factor acting either as activator or repressor of NOTCH1 pathway when respectively bound to NICD or histone deacetylases (HDACs). Compared with controls, NICD-mut transfectants had RBPJ preferentially complexed to NICD and showed higher levels of HDACs interacting with the promoter of the CD20 gene. Finally, treatment with the HDAC inhibitor valproic acid upregulated CD20 in both NICD-mut transfectants and primary CLL cells. In conclusion, NOTCH1 mutations are associated with low CD20 levels in CLL and are responsible for a dysregulation of HDAC-mediated epigenetic repression of CD20 expression.

Results of a randomized trial comparing high-dose chemotherapy plus Auto-SCT and R-FC in CLL at diagnosis.

The importance of early therapy intensification in B-cell CLL (B-CLL) patients remains to be defined. Even though several studies have been published, no randomized trials comparing directly autologous stem cell transplant (ASCT) and the accepted conventional therapy (that is, rituximab, fludarabine and CY; R-FC) have been reported so far. To assess the benefit of a first-line aggressive therapy, we designed a multicenter, randomized, phase 3 trial comparing R-FC and high-dose chemotherapy supported by ASCT in patients under 65 years of age, with stage B(II) or C B-CLL. Primary end point was CR: 96 patients were enrolled (48 in each arm). On an intent-to-treat basis, the CR rates in the ASCT and R-FC arms were 62.5% and 58%, respectively. After 5 years of follow-up, PFS was 60.4% in the ASCT arm and 65.1% in the R-FC arm, time to progression 65.8 and 70.5%, and overall survival 88% vs 88.1%, respectively. Our trial demonstrates, for the first time in a randomized manner, that frontline ASCT does not translate into a survival advantage when compared with benchmark chemoimmunotherapy in B-CLL patients; the possibility of its clinical benefit in certain subgroups remains uncertain.

Bendamustine in combination with ofatumumab in relapsed or refractory chronic lymphocytic leukemia: a GIMEMA Multicenter Phase II Trial.

We conducted a phase II, noncomparative, open-label, multicenter GIMEMA (Gruppo Italiano Malattie EMatologiche dell'Adulto) study (CLL0809) to assess the efficacy and safety of bendamustine in combination with ofatumumab (BendOfa) in relapsed/refractory chronic lymphocytic leukemia (CLL). Forty-seven patients from 14 centers were evaluated. Therapy consisted of bendamustine (70 mg/m(2)) for 2 consecutive days every 28 days, and ofatumumab 300 mg on day 1 and 1000 mg on day 8 during the first cycle, and 1000 mg on day 1 subsequently. Treatment was administered up to six cycles. The overall response rate (ORR), as per intention-to-treat analysis, was 72.3% (95% confidence of interval (CI), 57-84%), with 17% complete responses. After a median follow-up of 24.2 months, the overall survival was 83.6% (95% CI, 73.0-95.7%) and the progression-free survival (PFS) was 49.6% (95% CI, 35.9-68.6%). The median PFS was 23.6 months. Univariate and multivariate analyses were used to identify clinical and biological characteristics associated with ORR and PFS. Myelosuppression was the most common toxicity; grade ≥3 neutropenia was observed in 61.7% of patients; however, grade ≥3 infections occurred in 6% of patients. BendOfa is feasible and effective in relapsed/refractory CLL patients, including patients with high-risk clinical and biological features.

Intravenous injection of bortezomib, melphalan and dexamethasone in refractory and relapsed multiple myeloma.

BACKGROUND: A combination of bortezomib (1.3 mg/m(2)), melphalan (5 mg/m(2)), and dexamethasone (40 mg) (BMD), with all three drugs given as a contemporary intravenous administration, was retrospectively evaluated. PATIENTS AND METHODS: Fifty previously treated (median 2 previous lines) patients with myeloma (33 relapsed and 17 refractory) were assessed. The first 19 patients were treated with a twice-a-week (days 1, 4, 8, 11, 'base' schedule) administration while, in the remaining 31 patients, the three drugs were administered once a week (days 1, 8, 15, 22, 'weekly' schedule). RESULTS: Side-effects were predictable and manageable, with prominent haematological toxicity, and a better toxic profile in 'weekly' schedule (36% versus 66% in 'base' schedule). The overall response rate was 62%. After median follow-up of 24.5 months (range 2.7-50 months), the median progression-free survival (PFS) was 21.6 with no difference between the two schedules and the median overall survival (OS) was 33.8 months. Independently from the adopted schedule, we found that also in a cohort of relapsed/refractory patients achieving at least partial remission improved PFS (35.2 versus 9 months) and OS (unreached median versus 18 months). CONCLUSION: Taken together, our observations suggest that BMD is an effective regimen in advanced myeloma patients with acceptable toxicity.

BRIT1/MCPH1 expression in chronic myeloid leukemia and its regulation of the G2/M checkpoint.

BRIT1 (BRCT-repeat inhibitor of hTERT expression), also known as microcephalin (MCPH1), is a crucial gene in the complex cellular machine that is devoted to DNA repair and acts as a regulator of both the intra-S and G2/M checkpoints. The most important role of BRIT1/MCPH1 in the regulation of cell cycle progression appears to be the G2/M checkpoint. The K562 and peripheral blood cells of chronic myeloid leukemia (CML) patients at diagnosis were found to downregulate BRIT1/MCPH1. However, we could not find any correlation between bcr/abl activity and the BRIT1/MCPH1 level. In order to study the genomic instability of CML cells, we evaluated the ability of these cells to arrest mitotic division after exposure to hydroxyurea, a known genotoxic agent. We showed that CML cells continue to proliferate without the activation of the G2/M cell cycle checkpoint arrest or of the apoptotic mechanism. This behavior may predispose the cells to accumulate genomic defects. In conclusion, we found that CML cells have a low BRIT1/MCPH1 level and show a defective G2/M arrest, confirming that these cells have a constitutive genomic instability.

Is endoscopic ultrasound clinically useful for follow-up of gastric lymphoma?

BACKGROUND: Endoscopic ultrasound (EUS) is considered the best technique for locoregional staging at diagnosis but its role in the follow-up of patients with gastric lymphoma after organ-conserving strategies has not been established. DESIGN AND METHODS: We retrospectively evaluated 23 patients with primary gastric lymphoma treated with a stomach-conservative approach. Sixteen of them were affected by MALT lymphoma and seven by diffuse large-B-cell lymphoma (DLBCL). Five patients were treated with Helicobacter pylori (HP) eradication therapy alone (omeprazole + amoxicillin + clarithromycin); eight patients received a treatment including HP eradication and chemotherapy and the remaining 10 patients were treated with chemotherapy alone. RESULTS: At the end of treatment, a complete remission was documented in 21 (91%) patients by endoscopy with biopsy (E-Bx) but in only seven (30%) patients by EUS. A total of 99 evaluations with both EUS and E-Bx were evaluated and we found concordance between the two methods in 33 occasions (33%) only. No significant difference on the percentage of concordance was recorded between MALT and DLBCL. After a median follow-up of 36.5 months we have not observed any relapse in 12 patients (six DLBCL and six MALT) with a persistent positive EUS but negative E-Bx. CONCLUSIONS: Although the length of follow-up cannot exclude late relapse, we think that in restaging and follow-up of gastric lymphoma, EUS seems not to be a reliable tool if it is abnormal and E-Bx still remains the gold standard. Therefore, after conventional conservative treatment, persistence of EUS abnormality with a negative histology should not be considered as a clinically relevant persistence of disease and should not be a reason for further treatment.

NF-kB inhibition as a strategy to enhance etoposide-induced apoptosis in K562 cell line.

NF-kB is a transcription factor that mediates antiapoptotic signals in several cancer cell lines. Here we have demonstrated that the cytotoxic drug, Etoposide, activates NF-kB in K562, a chronic myeloid leukemia blast crisis cell line. Treatment with the NF-kB inhibitors MG-132, Bay11-7082, and Resveratrol impedes Etoposide-induced NF-kB activation, rendering K562 sensitive to Etoposide-induced apoptosis. Stable expression of mutant form of IkB-alpha, which retains NF-kB inactive in the cytoplasm of cells, confirmed the data obtained with molecular inhibitors. Both inhibitors and stable expression of SR-IkB are associated with down-modulation of the antiapoptotic protein Bcl-xL, suggesting that the survival pathway activated by Etoposide involves NF-kB-mediated Bcl-xL expression.

Protein free diet feeding: effects on sympathetic activity and salivary evoked secretion in the submandibular gland of the rat.

Protein restriction impairs the salivary flow rate and composition in human and rats. The aim of the present work was to establish the effect of low protein (casein 5%) and protein free (casein 0%) isocaloric diets on sympathetic activity and salivary evoked secretion in the submandibular gland (SMG) of the rat. After 21 days, rats fed casein 0% presented: (a) a significant shift to the left of the dose-response curves (DRC) to the autonomic agonists-norepinephrine (NE), methoxamine, isoproterenol (ISO) and methacholine; (b) increased food consumption (p<0.001); (c) decreased body (p<0.001) and SMG (p<0.001) weights maintaining SMG/body (w/w) relation; (d) enhanced submandibular alpha1-adrenoceptor number without changes in the apparent dissociation constant (Kd); (e) increased submandibular NE content (p<0.05) and phosphoinositoside hydrolysis (p<0.001); (f) decreased submandibular tyrosine hydroxylase activity (TH) (p<0.01). Casein 5% feeding increased food consumption (p<0.01) and reduced body weight (p<0.05). This protein restriction increased metacholine-evoked salivation, but it altered neither submandibular sympathetic activity nor sympathetic-induced salivary secretion as compared to the Control group (C) fed a similar diet containing 25.5% protein. Present results suggest that in the adult rat, a protein free diet during 21 days lowers SMG sympathetic and cholinergic activity leading to supersensitivity as revealed by up-regulation of alpha1-adrenergic receptor number and increased autonomic-evoked salivation.

Clinical and endoscopic presentation of primary gastric lymphoma: a multicentre study.

BACKGROUND: Although the stomach is the most frequent site of intestinal lymphomas, few data are available on both clinical endoscopic presentation of gastric lymphoma and possible differences between low-grade and high-grade lymphomas. METHODS: Clinical, histological and endoscopic records of consecutive patients with primary low-grade or high-grade lymphoma diagnosed were retrieved. Symptoms were categorized as 'alarm' or 'not alarm'. The endoscopic findings were classified as 'normal' or 'abnormal'. RESULTS: Overall, 144 patients with primary gastric lymphoma were detected, including 74 low-grade and 70 high-grade lymphoma. Alarm symptoms, particularly persistent vomiting and weight loss, were more frequently present in patients with high-grade lymphoma than in those with low-grade lymphoma (54% vs. 28%; P = 0.002). Low-grade lymphomas presented as 'normal' appearing mucosa (20% vs. 0%; P = 0.0004) or petechial haemorrhage in the fundus (9% vs. 0%; P = 0.02) more frequently than high-grade lymphomas, being also more often confined to the antrum (47% vs. 27%, P = 0.03) and associated with Helicobacter pylori infection (88% vs. 52%, P < 0.0001). On the contrary, high-grade lymphomas presented more commonly as ulcerative type (70% vs. 52%; P = 0.03), being also more frequently diagnosed in stage >I when compared with low-grade lymphomas (70% vs. 21%, P < 0.0001). CONCLUSIONS: The overall prevalence of alarm symptoms is quite low and may be absent in more than 70% of patients with low-grade lymphoma.

Divergent effects of corticotropin releasing hormone on endothelial cell nitric oxide synthase are associated with different expression of CRH type 1 and 2 receptors.

1. Endothelium is a target for an array of factors involved in inflammation. Endothelial cells express receptors for CRH, a neuropeptide produced during inflammation. We report both the concentration-dependent inhibitory effect of CRH upon cytokine-stimulated nitrite release by H5V murine endothelioma cells, and its stimulatory one in HUVEC cells. 2. Western blot analysis showed that CRH inhibits cytokine-stimulated iNOS protein in H5V cells, and, instead, potentiated it in HUVEC cells. 3. H5V cells expressed both CRH receptors (CRH-R1 and R2) mRNAs, whereas HUVEC cells expressed the CRH-R2 mRNA solely. 4. CRH increased medium nitrites and iNOS protein expression in H5V cells pretreated with the selective CRH-R1 antagonist CP 154,526. However, the selective CRH-R2 antagonist anti-Svg-30 failed to produce similar effects. In fact, anti-Svg-30 inhibited CRH-induced increase of nitrite release and iNOS expression in HUVEC cells. 5. Our results confirm the activating role of CRH on endothelial cells, although it suggests its possible inhibitory role in the late phase of the inflammatory response. NO-mediated effects of CRH on endothelial cells could be exploited in therapeutic strategies related to inflammatory and/or degenerative diseases.

Daily changes in beta-adrenergic sensitivity of rat submandibular gland. Correlation with beta-adrenoceptor rhythm.

Neurons from the superior cervical ganglia (SCG) innervate the submandibular gland and release noradrenaline during the dark phase of the daily photoperiod. Since in the pineal, another structure innervated by sympathetic neurons, nocturnal activation of the SCG is associated with beta-adrenergic sub- and super-sensitivity rhythms, the possible existence of similar phenomena in the rat submandibular gland was assessed. Wistar female rats, kept on a 14:10 light/dark cycle (light from 06:00 to 20:00 h), were sacrificed at 09:00, 14:00, 20:00, 24:00 and 04:00 h. beta-Adrenoceptors were studied by 3H-dihydroalprenolol binding to membrane preparations. The equilibrium dissociation constant (Kd) did not change as a function of time while significant daily variations in maximal binding values (Bmax) were observed with a peak at 20:00 h. Changes in Bmax correlated with a high response of adenylate cyclase to isoproterenol. In addition, when the response in salivary flow to isoproterenol was measured. a shift to the left (about 1 logarithmic unit) in dose-response curves was observed at 19:00-20:00 has compared to 08:00-09:00 h. These daily variations in isoproterenol responsiveness seem not to depend on the pattern of eating since a 24-h starvation or a nocturnal starvation for 16-18 days did not abolish the morning-evening differences in the salivary flow response to isoproterenol. Rather, the results suggest that the daily variations in isoproterenol response correlate with beta-adrenergic super- and sub-sensitivity phenomena associated with the circadian release of noradrenaline from SCG neurons.

Tamoxifen inhibits nerve growth factor-induced proliferation of the human breast cancerous cell line MCF-7.

An array of polypeptide growth factors contribute to the development of breast cancer, the most common tumor-related cause of death in women of Western countries. Therefore, breast cancer therapy should be aimed at inhibition of growth factor-dependent breast cancerous cell proliferation. However, the relative contribution of each individual factor in the development and maintenance of the transformed phenotype is largely unknown. Here we report for the first time that the proliferative effects of nerve growth factor, (NGF) a typical neurotrophin, are similar to those of epidermal growth factor (EGF) and insulin-like growth factor II, and are enhanced by 17beta-estradiol in the human breast cancer cell line MCF-7. The effect of NGF appeared to be mediated by its trkA receptors (trkA(NGFR)), as suggested by the potent inhibition of both MCF-7 cell proliferation and trkA(NGFR) phosphorylation occurring upon treatment of cultures with the selective trkA(NGFR) inhibitor K252a. Surprisingly, the antiestrogen drug tamoxifen (TAM) inhibited NGF-induced MCF-7 cell proliferation and trkA(NGFR) phosphorylation in a concentration-related fashion. The effect of TAM seemed to be estrogen receptor-independent, because the pure estrogen receptor antagonist ICI 182.780 was unable to block NGF-induced trkA(NGFR) phosphorylation. Our data underline the new emerging role of trkA(NGFR) in breast tumor growth, and suggest a related novel therapeutic use of TAM in breast cancer.

Increased ACTH and cortisol secretion after interleukin-alpha injection in the common marmoset (Callithrix jacchus jacchus).

We have studied the effect of intravenous injection of interleukin-1 (dose range: from 0.25 to 4.5 microg/kg of body weight) on plasma ACTH and cortisol levels in the marmoset, a primate paradygm of peripheral glucocorticoid resistance. Blood sampling were collected and body temperature recorded 0, 15, 30, 60, 120, 180, 240 and 300 min after injection. Interleukin-1 stimulated secretion of ACTH in a dose-dependent fashion. Maximal secretion occurred 120 min after injection, and lasted up to 240 min. Plasma ACTH levels returned to baseline 300 min after interleukin-1 injection. Plasma cortisol levels were related to ACTH levels. Body temperature elevation, which occurred 10-15 min after injection was dose-dependent, and lasted 3 h. Results suggest that the pyrogenic effect of interleukin is associated, in the marmoset, with integrated activation of the hypothalamic-pituitary-adrenal axis. In light of the proneness of marmosets to hyperimmune disorders, our data are consistent with the hypothesized central biological role of IL-1, as well as the pathophysiological relevance of the neuro-endocrine-immune cross-talk during the acute phase response.

Apomorphine, dopamine and phenylethylamine reduce the proportion of phosphorylated insulin receptor substrate 1.

We tested the ability of dopamine, apomorphine, phenylethylamine and pergolide to inhibit the proliferation of fetal calf serum-stimulated human breast cancer (MCF)-7 cells. While the first three compounds were able to block the proliferation of MCF-7 cells, pergolide failed to do so (up to 100 microM). The inhibitory effect of dopamine, apomorphine and phenylethylamine was also evident in serum-starved insulin-stimulated MCF-7 cells. Apomorphine also inhibited the proliferation of the human oestrogen receptor-negative breast cancer (MDA-MB231) and prostate carcinoma (LNCaP) cell lines. In a second set of experiments, we measured the ability of dopamine, apomorphine, phenylethylamine and pergolide to inhibit the phosphorylation (or increase the dephosphorylation) of the insulin receptor substrate (IRS)-1, a major intracellular substrate of the insulin-like growth factor (IGF)-1 receptor. Dopamine, apomorphine and phenylethylamine all reduced to zero the level of phosphorylated IRS-1 with potencies ranging between 0.01 and 1 microM. Finally, we found that fibroblasts from IRS-1 null (-/-) mice were less sensitive to the anti-proliferative effect of apomorphine compared to fibroblasts from wild type-mice, suggesting that the inhibition of IRS-1 phosphorylation by apomorphine is an important aspect of the activity of this compound.

The effects of epidural analgesia and uterine contractions on fetal oxygen saturation during the first stage of labour.

The aim of this study was to investigate the effects of (1) maternal epidural analgesia and (2) uterine contractions on fetal oxygen saturation. After informed consent 18 women were included in our prospective, non-blinded, observational study. After 30 min of monitoring fetal oxygen saturation and uterine contractions, all the parturients, at cervical dilatation >/= 3 cm, received epidural analgesia for labour: a Pajunk epidural catheter was passed through a 17-gauge Tuohy needle and left in the lumbar epidural space (insertion level L3-4 or L2-3). Sufentanil 10 microg and 0.1% ropivacaine 15 mL were injected into the epidural catheter. A second and third 15-mL epidural dose of 0.1% ropivacaine were administered on patient demand. Fetal oxygen saturation was unaffected by epidural analgesia and there was no change in fetal SpO(2)following an uncomplicated epidural top-up. SpO(2)values (%) 30 s before, during and after contraction were 47.6 +/- 2.4, 52.5 +/- 5.3 and 42.9 +/- 7.2 respectively. These changes were significant. A contraction appeared to inject a bolus of oxygenated blood into the fetus, causing an initial increase in fetal oxygenation followed by a decline. The lowest SpO(2)values observed occurred 120 s after the start of contractions.

Mitogenic effect of nerve growth factor (NGF) in LNCaP prostate adenocarcinoma cells: role of the high- and low-affinity NGF receptors.

We have investigated the effect of nerve growth factor (NGF) in the androgen-dependent, prostate adenocarcinoma LNCaP cell line. Exposure of LNCaP cells to NGF resulted in a significant increase of cell proliferation. The effect was concentration dependent and equally present in serum- or charcoal-stripped serum-supplemented and serum-deprived conditions. The mitogenic action of NGF was accompanied by an enhanced expression of prostate-specific antigen (PSA) and resulted additive to the proliferative effect of dihydrotestosterone. The proliferative effect of NGF appeared to be mediated by the high-affinity NGF receptor, p140trka. Only p140trka, but not the low-affinity NGF receptor, p75LNGFR, was expressed in LNCaP cells; both the proliferative response and the phosphorylation of p140trka upon NGF treatment were prevented by the tyrosine kinase inhibitor K252a. LNCaP cells transiently transfected with the cDNA encoding for p75LNGFR appeared more sensitive to NGF, as demonstrated by the increased number of p75LNGFR-transfected LNCaP cells exposed for 72 h to NGF compared with wild LNCaP cultures. However, p75LNGFR-transfected LNCaP cells rapidly underwent apoptotic death when deprived of NGF. Our study demonstrates the physiological relevance of NGF in the regulation of prostate cell proliferation and the relative contribution of the high- and low-affinity NGF receptors in this control.

Responsiveness of irradiated rat anterior pituitary cells to hypothalamic releasing hormones is restored by treatment with growth hormone.

Hypopituitarism is a common sequela of irradiation in cancer patients. Here we report that recombinant human growth hormone (r-hGH) prevents cell death and restores secretory capacity of irradiated rat pituitary cells in vitro. Dispersed rat pituitary cells from male Sprague-Dawley rats, irradiated with a 9-Gy sublethal dose, were incubated with r-hGH before, after, or before and after irradiation. Treatment with GH resulted in increased cell survival, which reached its maximum at the concentration of 5 nM, with an EC(50) of 3.5 nM. Protective effects of GH on pituitary cells were more pronounced in cultures treated before and after irradiation. Similarly, beneficial effects of GH were observed on the secretory capacity of surviving cells. In fact, irradiated pituitary cells treated with GH secreted substantial amounts of GH, luteinizing hormone, follicle-stimulating hormone, prolactin, thyroid-stimulating hormone and adrenocorticotropic hormone in response to specific releasing hormones. Such effects of GH were prevented in the presence of the specific GH receptor antagonists B2036 and G120K. Our results show that r-hGH exerts a specific protective effect on irradiated rat pituitary cells and suggest possible use of GH as an adjuvant agent for prevention of postirradiation hypopituitarism.