Protocol of a clinical trial on the effectiveness of pudendal nerve block with and without neurostimulation in reducing posthemorroidectomy pain.

INTRODUCTION: Hemorrhoidal pathology is the most frequent proctological problem with a prevalence of 44% of the adult population. The most effective treatment is surgery but it also has the highest postoperative pain rate with moderate to severe pain rates of 30-40% during the first 24-48 hours. Here lies the importance of seeking measures to improve this situation, such as the pudendal nerve block with local anesthetic. However, the variability of the pudendal nerve sometimes makes its blockade ineffective and for this reason nerve location methods are sought to achieve a higher rate of success. The main aim of the study is to compare pain in the immediate postoperative period (24 h) after hemorrhoidectomy in patients with pudendal nerve block guided by anatomical references and guided by neurostimulation. METHODS: The present project proposes the performance of a single-center, triple-blind, randomized clinical trial of efficacy, carried out under conditions of routine clinical practice. Patients over 18 years old with hemorrhoids refractory to medical treatment, symptomatic grade III-IV and grade II hemorrhoids that do not respond to conservative procedures in a third level hospital in Spain and that are subsidiaries of surgery in major ambulatory surgery will be included. Demographic variables, variables on hemorrhoidal pathology, details of surgery, verbal numeric pain scale in the preoperative period and surgical complications will be collected. RESULTS: Not avaliable until the end of the study. CONCLUSIONS: The pudendal nerve block guided by anatomical landmarks has been shown to be useful in postoperative pain control after hemorrhoidectomy although the use of the neurostimulator has not been well studied and we believe it may improve outcom.

Long-Term Safety of In Utero Exposure to Anti-TNFα Drugs for the Treatment of Inflammatory Bowel Disease: Results from the Multicenter European TEDDY Study.

OBJECTIVES: The long-term safety of exposure to anti-tumor necrosis factor (anti-TNFα) drugs during pregnancy has received little attention. We aimed to compare the relative risk of severe infections in children of mothers with inflammatory bowel disease (IBD) who were exposed to anti-TNFα drugs in utero with that of children who were not exposed to the drugs. METHODS: Retrospective multicenter cohort study. Exposed cohort: children from mothers with IBD receiving anti-TNFα medication (with or without thiopurines) at any time during pregnancy or during the 3 months before conception. Non-exposed cohort: children from mothers with IBD not treated with anti-TNFα agents or thiopurines at any time during pregnancy or the 3 months before conception. The cumulative incidence of severe infections after birth was estimated using Kaplan-Meier curves, which were compared using the log-rank test. Cox-regression analysis was performed to identify potential predictive factors for severe infections in the offspring. RESULTS: The study population comprised 841 children, of whom 388 (46%) had been exposed to anti-TNFα agents. Median follow-up after delivery was 47 months in the exposed group and 68 months in the non-exposed group. Both univariate and multivariate analysis showed the incidence rate of severe infections to be similar in non-exposed and exposed children (1.6% vs. 2.8% per person-year, hazard ratio 1.2 (95% confidence interval 0.8-1.8)). In the multivariate analysis, preterm delivery was the only variable associated with a higher risk of severe infection (2.5% (1.5-4.3)). CONCLUSIONS: In utero exposure to anti-TNFα drugs does not seem to be associated with increased short-term or long-term risk of severe infections in children.

Novel clinical and molecular findings in Spanish patients with naevoid basal cell carcinoma syndrome.

BACKGROUND: Naevoid basal cell carcinoma syndrome (NBCCS) is an autosomal dominant disorder characterized by developmental alterations and multiple basal cell carcinomas. Mutations in PTCH1, which encodes a membrane receptor for Sonic Hedgehog, are associated with the development of the disease. Most of them produce a truncated protein, which is unable to suppress Smoothened protein and continuously activates the downstream pathway. OBJECTIVES: We aimed to characterize 22 unrelated Spanish patients with NBCCS, the largest cohort with Gorlin syndrome reported to date in Spain. METHODS: Genomic analysis of PTCH1 was performed in patients with NBCCS and controls, and mutations were analysed using bioinformatics tools. RESULTS: We report for the first time two young patients, one each with uterus didelphys and ganglioneuroma, within the context of NBCCS. One patient showing a severe phenotype of the disease had developed basal cell carcinomas since childhood. Sanger sequencing of PTCH1 in this cohort identified 17 novel truncating mutations (11 frameshift, five nonsense and one mutation affecting an exon-intron splice site) and two novel missense mutations that were predicted to be pathogenic. The patients showed great clinical variability and inconsistent genotype-phenotype correlation, as seen in relatives carrying similar mutations. CONCLUSIONS: This study contributes to increase the pool of clinical manifestations of NBCCS, as well as increasing the number of pathogenic mutations identified in PTCH1 predisposing to the condition. The inconsistencies found between phenotype and genotype suggest the involvement of other modifying factors, genetic, epigenetic or environmental.

IL-11 and CCL-1: Novel Protein Diagnostic Biomarkers of Lung Adenocarcinoma in Bronchoalveolar Lavage Fluid (BALF).

INTRODUCTION: Lung cancer (LC) and chronic obstructive pulmonary disease (COPD) are smoking-related diseases, with the presence of COPD itself increasing the risk for development of LC, probably owing to underlying inflammation. LC is typically detected at late stages of the disease and carries a poor prognosis. There is an unmet need for methods to facilitate the early detection of LC in high-risk subjects such as smokers. METHODS: The expression of inflammatory proteins in bronchoalveolar lavage fluid (BALF) samples was studied by antibody arrays in a prospective cohort of 60 smokers of more than 30 pack-years divided into four groups (control, patients with LC, patients with COPD, and patients with LC plus COPD). Relevant biomarkers were validated by Western blot. Additional validation with enzyme-linked immunosorbent assay (ELISA) was carried out on two independent controlled cohorts of 139 patients (control, patients with LC, patients with COPD, and patients with LC plus COPD) and 160 patients (control and patients with LC of all histological types). RESULTS: A total of 16 differentially expressed proteins in samples from patients with LC, COPD, and LC plus COPD were identified by antibody arrays and validated by Western blot and ELISA. C-C motif chemokine ligand 1 (CCL-1) and interleukin-11 (IL)-11 were selectively expressed in samples from patients with adenocarcinoma with or without COPD (p < 0.005). These proteins exhibited a remarkable diagnostic performance for lung adenocarcinoma in an independent cohort of 139 patients. Receiver operating characteristic curves showed that the optimum diagnostic cutoff value for IL-11 was 42 pg/mL (area under the curve = 0.93 [95% confidence interval: 0.896-0.975], sensitivity 90%, specificity 86%), whereas for CCL-1 it was 39.5 pg/mL (0.83 [95% confidence interval: 0.749-0.902], sensitivity 83%, and specificity 74%). Further validation of the ELISA biomarkers at the aforementioned cutoffs was performed in an additional cohort of 160 patients (20 controls, 66 patients with LC, and 74 patients with LC plus COPD). There was a significant correlation between BALF levels of IL-11 and CCL-1 (r2 = 0.76, p < 0.001), and the use of both biomarkers increased the diagnostic accuracy to 96.1% in the two validation cohorts. Appropriate diagnostic performance was observed for all subgroups regardless of stage at diagnosis, involvement of the bronchial tract, pack-years smoked, and number of cells in BALF. CONCLUSIONS: IL-11 and CCL-1 are highly specific biomarkers with great accuracy for the diagnosis of lung adenocarcinoma in BALF specimens. Further study of these proteins as markers for the early diagnosis and screening of plasma and other biological materials is warranted.

Loss of the tumor suppressor spinophilin (PPP1R9B) increases the cancer stem cell population in breast tumors.

The spinophilin (Spn, PPP1R9B) gene is located at 17q21.33, a region frequently associated with microsatellite instability and loss of heterozygosity, especially in breast tumors. Spn is a regulatory subunit of phosphatase1a (PP1), which targets the catalytic subunit to distinct subcellular locations. Spn downregulation reduces PPP1CA activity against the retinoblastoma protein, pRb, thereby maintaining higher levels of phosphorylated pRb. This effect contributes to an increase in the tumorigenic properties of cells in certain contexts. Here, we explored the mechanism of how Spn downregulation contributes to the malignant phenotype and poor prognosis in breast tumors and found an increase in the stemness phenotype. Analysis of human breast tumors showed that Spn mRNA and protein are reduced or lost in 15% of carcinomas, correlating with a worse prognosis, a more aggressive tumor phenotype and triple-negative tumors, whereas luminal tumors showed high Spn levels. Downregulation of Spn by shRNA increased the stemness properties along with the expression of stem-related genes (Sox2, KLF4, Nanog and OCT4), whereas ectopic overexpression of Spn cDNA reduced these properties. Breast tumor stem cells appeared to have low levels of Spn mRNA, and Spn loss correlated with increased stem-like cell appearance in breast tumors as indicated by an increase in CD44+/CD24- cells. A reduction of the levels of PPP1CA mimicked the cancer stem-like cell phenotype of Spn downregulation, suggesting that the mechanism of Spn involves PP1a. These increased cancer stem cell-like properties with reduced Spn might account for the malignant phenotype observed in Spn-loss tumors and may contribute to a worse patient prognosis.

Aß promotes VDAC1 channel dephosphorylation in neuronal lipid rafts. Relevance to the mechanisms of neurotoxicity in Alzheimer's disease.

Voltage-dependent anion channel (VDAC) is a mitochondrial protein abundantly found in neuronal lipid rafts. In these membrane domains, VDAC is associated with a complex of signaling proteins that trigger neuroprotective responses. Loss of lipid raft integrity may result in disruption of multicomplex association and alteration of signaling responses that may ultimately promote VDAC activation. Some data have demonstrated that VDAC at the neuronal membrane may be involved in the mechanisms of amyloid beta (Aß)-induced neurotoxicity, through yet unknown mechanisms. Aß is generated from amyloid precursor protein (APP), and is released to the extracellular space where it may undergo self-aggregation. Aß aggregate deposition in the form of senile plaques may lead to Alzheimer's disease (AD) neuropathology, although other pathological hallmarks (such as hyper-phosphorylated Tau deposition) also participate in this neurodegenerative process. The present study demonstrates that VDAC1 associates with APP and Aß in lipid rafts of neurons. Interaction of VDAC1 with APP was observed in lipid rafts from the frontal and entorhinal cortex of human brains affected by AD at early stages (I-IV/0-B of Braak and Braak). Furthermore, Aß exposure enhanced the dephosphorylation of VDAC1 that correlated with cell death. Both effects were reverted in the presence of tyrosine phosphatase inhibitors. VDAC1 dephosphorylation was corroborated in lipid rafts of AD brains. These results demonstrate that Aß is involved in alterations of the phosphorylation state of VDAC in neuronal lipid rafts. Modulation of this channel may contribute to the development and progression of AD pathology.

Effect of poly(propylene imine) glycodendrimers on ß-amyloid aggregation in vitro and in APP/PS1 transgenic mice, as a model of brain amyloid deposition and Alzheimer's disease.

Poly(propylene imine) (PPI) glycodendrimers are promising candidates as drug carriers and antiamyloidogenic and antiprionic agents. In this study the anti-ß-amyloid capacity of PPI glycodendrimers of the fourth and fifth generations was investigated in vitro and in vivo. We assessed distinct PPI glycodendrimers including G4mDS and G5mDS, with electroneutral maltose shell, and G4mOS and G4m-IIIOS, with cationic maltose or maltotriose shell. Our results show that in vitro PPI maltose dendrimers reduce the toxicity of Aß(1-42). However, only the electroneutral maltose dendrimers G4mDS and G5mDS reduce the toxicity of Alzheimer's disease brain extracts in SH-SY5Y neuroblastoma cells. PPI maltose dendrimers with electroneutral or cationic surface penetrate the cytoplasm of cultured cells, and they reach the brain when administered intranasally. Both cationic G4mOS and electroneutral G4mDS are able to modify the total burden of ß-amyloid in APP/PS1 mice. The studied dendrimers did not reverse memory impairment in APP/PS1 mice following chronic administration; moreover, cationic G4mOS caused cognitive decline in nontransgenic mice. In spite of the capacity of G4mDS and G4mOS to cross the blood-brain barrier and modulate Aß aggregation in APP/PS1 mice, further studies are needed to learn how to reduce the harmful effects of maltose dendrimers in vivo.

p38α limits the contribution of MAP17 to cancer progression in breast tumors.

MAP17 is a small, 17-kDa, non-glycosylated membrane protein that is overexpressed in a percentage of carcinomas. In the present work, we have analyzed the role of MAP17 expression during mammary cancer progression. We have found that MAP17 is expressed in 60% human mammary tumors while it is not expressed in normal or benign neoplasias. MAP17 levels increased with breast tumor stage and were strongly correlated with mammary tumoral progression. A significant increase in the levels of reactive oxygen species (ROS) was observed in MAP17-expressing cells, as compared with parental cells. This increase was further paralleled by an increase in the tumorigenic capacity of carcinoma cells but not in immortal non-tumoral breast epithelial cells, which provides a selective advantage once tumorigenesis has begun. Expression of specific MAP17 shRNA in protein-expressing tumor cells reduced their tumorigenic capabilities, which suggests that this effect is dependent upon MAP17 protein expression. Our data show that ROS functions as a second messenger that enhances tumoral properties, which are inhibited in non-tumoral cells. We have found that p38α activation mediates this response. MAP17 triggers a ROS-dependent, senescence-like response that is abolished in the absence of p38a activation. Furthermore, in human breast tumors, MAP17 activation is correlated with a lack of phosphorylation of p38α. Therefore, MAP17 is overexpressed in late-stage breast tumors, in which oncogenic activity relies on p38 insensitivity to induce intracellular ROS.

α-synuclein phosphorylation and truncation are normal events in the adult human brain.

α-synuclein is a key protein in Lewy body diseases (LBDs) and a major component of Lewy bodies and related aberrant cytoplasmic and neuritic inclusions. Regional differences in α-synuclein have been associated with selective neuronal vulnerability to Lewy pathology. Furthermore, phosphorylation at serine 129 (Ser129) and α-synuclein truncation have been considered crucial in the pathogenesis of Lewy inclusions. The present study shows consistent reduction in α-synuclein protein expression levels in the human substantia nigra and nucleus basalis of Meynert compared with other brain regions independently of age and pathology. Phosphorylated α-synuclein at Ser129 is naturally increased in these same regions, thus inversely related with the total amount of α-synuclein. In contrast, truncated α-synuclein is naturally observed in control and diseased brains and correlating with the total amount of α-synuclein. Several truncated variants have been identified where some of these variants are truncated at the C-terminal domain, whereas others are truncated at the N-terminal domain, and all are present in cases with and without Lewy pathology. Although accumulation of truncated α-synuclein variants and phosphorylated α-synuclein occurs in Lewy bodies, α-synuclein phosphorylation and truncation can be considered constitutive in control and diseased brains.

Neuropathological changes correlate temporally but not spatially with selected neuromodulatory responses in natural scrapie.

AIM: Neuropathological changes classically associated with sheep scrapie do not always correlate with clinical disease. We aimed to determine if selected neuromodulatory responses were altered during the course of the infection as it has been described in Creutzfeldt-Jakob disease and experimental bovine spongiform encephalopathy. METHODS: Hemi-brains from healthy sheep and natural scrapie cases at two stages of infection were examined for biochemical alterations related to the expression of type I metabotropic glutamatergic receptors (mGluR(1) ) and type I adenosine receptors I (A(1) R), and of selected downstream intermediate signalling targets. Immunohistochemistry for different scrapie-related neuropathological changes was performed in the contralateral hemi-brains. RESULTS: PrP(d) deposition, spongiform change, astrocytosis and parvalbumin expression were significantly altered in brains from clinically affected sheep compared with preclinical cases and negative controls; the latter also showed significantly higher immunoreactivity for synaptophysin than clinical cases. Between clinically affected and healthy sheep, no differences were found in the protein levels of mGluR(1) , while phospholipase Cß1 expression in terminally ill sheep was increased in some brain areas but decreased in others. Adenyl cyclase 1 and A(1) R levels were significantly lower in various brain areas of affected sheep. No abnormal biochemical expression levels of these markers were found in preclinically infected sheep. CONCLUSIONS: These findings point towards an involvement of mGluR(1) and A(1) R downstream pathways in natural scrapie. While classical prion disease lesions and neuromodulatory responses converge in some affected regions, they do not do so in others suggesting that there are independent regulatory factors for distinct degenerative and neuroprotective responses.

B-cell translocation gene 2 is over-expressed in peri-infarct neurons after ischaemic stroke.

OBJECTIVES: Recovery from stroke is dependent on the survival of neurons in the dynamic peri-infarcted region. Although several markers of neuronal injury and apoptotic cell death have been described, administration of neuroprotective drugs directed at specific molecules has had limited success. A complete understanding of deregulated genes associated with neuronal death would be beneficial. Our previous microarray studies identified increased expression of a novel protein, the B-cell translocation gene 2 (BTG2), in infarcted regions. METHODS: We have used immunohistochemistry and Western blotting to examine the expression and localization of BTG2 in stroked brain tissue and immunofluorescent staining of human fetal brain neurons to determine if oxygen-glucose deprivation affected its expression. RESULTS: We show that BTG2 is strongly expressed in peri-infarcted and infarcted regions of brain tissue, localizing in neuronal nuclei and cytoplasm, whilst being absent or very weakly expressed in normal looking contralateral tissue. Exposure of human fetal brain neurons to oxygen-glucose deprivation also induced BTG2 expression in the cytoplasm and perinuclear regions of cells staining positive for propidium iodide (a marker of nuclear damage). CONCLUSIONS: BTG2 may be a modulator of cell survival and differentiation and could help to protect against cell death by inhibition of necrosis and/or apoptotic signalling pathways.

S-adenosylhomocysteine hydrolase downregulation contributes to tumorigenesis.

With the idea to discover novel genes involved in proliferation, we have performed a genome-wide loss-of-function genetic screen to identify additional putative tumor suppressor genes. We have previously identified five genes belonging to different biochemical families. In this report, we focused on the study of one of these genes designated S-adenosylhomocysteine hydrolase (SAHH), which has also been previously identified in an independent short hairpin RNA screening. SAHH inactivation confers resistance to both p53 and p16(INK4)-induced proliferation arrest. Interestingly, SAHH inactivation inhibits p53 transcriptional activity and impairs DNA damage-induced transcription of p21(Cip1). Given that SAHH downregulation modulates senescence in primary cells, we also studied SAHH expression in human tumors at the messenger RNA (mRNA) and protein levels. SAHH mRNA was lost in 50% of tumor tissues from 206 patients with different kinds of tumors in comparison with normal tissue counterparts. Moreover, SAHH protein was also affected in some colon cancers. Such findings may be of relevance to cancer research, suggesting that SAHH might be a largely unexplored tumor suppressor.

Modulation of SIRT1 expression in different neurodegenerative models and human pathologies.

We examined the expression of SIRT1 in several experimental paradigms of human pathologies. We used a neuroblastoma cell line (B65), neuronal primary cultures (hippocampus and cerebellar granule cells) and in vivo approaches in rat and senescence murine models (SAM). Cell cultures and rats were treated with several well-know neurotoxins, i.e. rotenone, MPP(+), kainate and 3-nitropropionic acid. Subsequently, SIRT1 expression was compared in these different paradigms of neurotoxicity. The pattern of expression of SIRT1 in proliferating cell cultures (B65) was different to that in quiescent cell cultures. In the murine model of senescence (senescence-accelerated mice prone, SAMP8), SIRT1 expression progressively decreased, while in the control strain (senescence-accelerated mice resistant, SAMR1) it increased. Finally, we studied human samples of Parkinson's disease (PD), dementia with Lewy bodies (DLB) and Huntington's diseases (HD). SIRT1 expression decreased dramatically in HD, but there were no significant changes in Parkinson-related illnesses. In conclusion, SIRT1 expression may be a good sensor of toxic neuronal processes.

Phosphorylation of tau and alpha-synuclein in synaptic-enriched fractions of the frontal cortex in Alzheimer's disease, and in Parkinson's disease and related alpha-synucleinopathies.

Phosphorylation of tau and phosphorylation of alpha-synuclein are crucial abnormalities in Alzheimer's disease (AD) and alpha-synucleinopathies (Parkinson's disease: PD, and dementia with Lewy bodies: DLB), respectively. The presence and distribution of phospho-tau were examined by sub-fractionation, gel electrophoresis and Western blotting in the frontal cortex of cases with AD at different stages of disease progression, PD, DLB pure form and common form, and in age-matched controls. Phospho-tauSer396 has been found in synaptic-enriched fractions in AD frontal cortex at entorhinal/transentorhinal, limbic and neocortical stages, thus indicating early tau phosphorylation at the synapses in AD before the occurrence of neurofibrillary tangles in the frontal cortex. Phospho-tauSer396 is also found in synaptic-enriched fractions in the frontal cortex in PD and DLB pure and common forms, thus indicating increased tau phosphorylation at the synapses in these alpha-synucleinopathies. Densitometric studies show between 20% and 40% phospho-tauSer396, in relation with tau-13, in synaptic-enriched fractions of the frontal cortex in AD stages I-III, and in PD and DLB. The percentage reaches about 95% in AD stage V and DLB common form. Yet tau phosphorylation characteristic of neurofibrillary tangles, as revealed with the AT8 antibody, is found in the synaptic fractions of the frontal cortex only at advanced stages of AD. Increased phosphorylated alpha-synucleinSer129 levels are observed in the synaptic-enriched fractions of the frontal cortex in PD and DLB pure and common forms, and in advanced stages of AD. Since tau-hyperphosphorylation has implications in microtubule assembly, and phosphorylation of alpha-synuclein at Ser129 favors alpha-synuclein aggregation, it can be suggested that synapses are targets of abnormal tau and alpha-synuclein phosphorylation in both groups of diseases. Tau phosphorylation at Ser396 has also been found in synaptic-enriched fractions in 12-month-old transgenic mice bearing the A53T alpha-synuclein mutation.

Identification of photocatalytic degradation products of bezafibrate in TiO(2) aqueous suspensions by liquid and gas chromatography.

In the present study the photocatalytic degradation of bezafibrate (BZF), a lipid regulator agent, has been investigated using TiO(2) suspensions and simulated solar light. The study focus on the identification of degradation products (DPs) using powerful analytical techniques such as liquid chromatography time of flight mass spectrometry (LC-TOF-MS), gas chromatography mass spectrometry (GC-MS), and high-performance liquid chromatography with diode-array detection (HPLC-DAD). Each technique provided complementary information that enabled the identification of 21 DPs. Accurate mass measurements obtained by LC-TOF-MS provided the elucidation of 17 DPs. Mass errors lower than 2mDa, allowed the assignment of empirical formula for the mayor DPs to be determined confidently. Three DPs were identified by GC-MS through the structural information provided by full scan mass spectra obtained by electron impact (EI) ionization and two more by HPLC-DAD by comparing the retention times (t(R)) and the UV spectra of the unknown DPs with those of commercial standards. Based on this by-product identification a possible multi-step degradation scheme was proposed. The pathways include single or multiple hydroxylation of BZF with subsequent phenoxy ring opening and the cleavage of the amide and ether bonds.

Expression of mutant ubiquitin (UBB+1) and p62 in myotilinopathies and desminopathies.

Protein aggregates in muscle cells are the morphological hallmark of myofibrillar myopathies, including myotilinopathies and desminopathies. The aim of the present study is to analyse the expression of mutant ubiquitin (UBB+1), an aberrant form of ubiquitin which accumulates in certain disorders characterized by intracellular aggregates of proteins, and p62, a multimeric signal protein which plays an active role in aggregate formation, in muscle biopsies from patients suffering from myotilinopathy and desminopathy in order to gain understanding of the mechanisms leading to protein aggregation in these disorders. Single immunohistochemistry, and single- and double-labelling immunofluorescence and confocal microscopy for UBB+1 and p62, has been performed in muscle biopsies from patients suffering from myotilinopathy and desminopathy. Strong UBB+1 immunoreactivity, colocalizing with myotilin aggregates, was found in muscle fibres in myotilinopathies. UBB+1 accumulation, colocalizing with desmin aggregates, also occurs in desminopathies. In addition, strong p62 immunoreactivity colocalizing with myotilin aggregates was observed in myotilinopathies. Similarly, p62 immunoreactivity colocalizing with desmin aggregates was found in desminopathies. The present findings suggest that accumulation of protein aggregates in myotilinopathies and in desminopathies may be related with UBB+1/abnormal protein complexes which are resistant to proteasome degradation. Furthermore, these observations suggest a relationship between the presence of p62 and the formation of inclusions in different subtypes of myofibrillar myopathies.

Expression levels of adenosine receptors in hippocampus and frontal cortex in argyrophilic grain disease.

Expression of adenosine receptors of the A1, A2A and A2B type has been examined in the post-mortem frontal cortex and hippocampus in argyrophilic grain disease (AGD), a tauopathy affecting the hippocampus but usually not the frontal cortex, in an attempt to learn about the modulation of the adenosine pathway in this disorder. Significant increased levels of A1, but not of A2A and A2B, have been observed in AGD in the hippocampus but not in the frontal cortex, when compared with age-matched controls. This is accompanied by increased levels of adenylyl cyclase (AC), an effector of A1, and by increased (although not significant) percentage of inhibition of forskolin-stimulated AC by the A1 agonist cyclohexyladenosine in the hippocampus in AGD. These findings indicate sensitization of A1/AC in the hippocampus in AGD, and support a putative activation of the A1/AC pathway that may facilitate protection of this preferentially involved region in AGD.

Expression of transcription factors CREB and c-Fos in the brains of terminal Creutzfeldt-Jakob disease cases.

Expression levels and localization of transcription factors cAMP response element binding protein (CREB(1) and CREB(2)) and c-Fos, as well as levels of up-stream mitogen-activated protein kinases/extracellular signal-regulated kinases (ERK-1 and ERK-2) and p38 kinase, were examined in the brains (frontal cortex) of eleven cases with Creutzfeldt-Jakob disease (CJD) and five age-matched controls. Preserved expression levels of ERK-1-P, ERK-2-P and p38-P were observed in CJD. However, significantly reduced levels, as revealed by gel electrophoresis and Western blotting, and reduced numbers of immunoreactive nuclei, as seen by immunohistochemistry, to CREB, CREB-P, c-Fos and c-Fos-P were found in CJD when compared with controls. These observations point to exhausted CREB and c-Fos brain responses, in spite of preserved up-stream signaling kinases, thus favoring cell death in terminal stages of CJD.