BRAF mutational status as a prognostic marker for survival in malignant melanoma: a systematic review and meta-analysis.

Background: The analysis of the BRAF mutational status has been established as a standard procedure during diagnosis of advanced malignant melanoma due to the fact that BRAF inhibitors constitute a cornerstone in the treatment of metastatic disease. However, the general impact of BRAF mutational status on survival remains unclear. Our study aimed to assess the underlying prognostic significance of BRAF mutant versus wild type (WT) malignant melanoma on overall survival (OS), disease-free survival (DFS) and progression-free survival (PFS).Material and methods: A systematic literature search in EMBASE, Medline and Cochrane CENTRAL was performed. Studies were included if they reported survival outcomes for BRAF mutant versus WT patients as hazard ratios (HR) or in Kaplan-Meier (KM) curves. Random-effects meta-analysis models were used to pool HRs across the studies.Results: Data from 52 studies, representing 7519 patients, were pooled for analysis of OS. The presence of a BRAF mutation was statistically significantly associated with a reduced OS (HR [95% confidence interval (CI)]: 1.23 [1.09-1.38]), however, with substantial heterogeneity between the studies (I2: 58.0%). Meta-regression and sensitivity analyses showed that age, sex and BRAF mutation testing method did not have a significant effect on the OS HR. BRAF mutant melanoma showed comparable effect on DFS to non-BRAF mutant melanoma in stage I-III melanoma (combined HR: 1.16, 95% CI: 0.92-1.46), and on PFS in stage III-IV (HR: 0.98 (95% CI: 0.68-1.40)).Conclusion: Although there was substantial heterogeneity between the studies, the overall results demonstrated a poorer prognosis and OS in patients harbouring BRAF mutations. Future studies should take this into account when evaluating epidemiological data and treatment effects of new interventions in patients with malignant melanoma.

Five-year follow-up of 11C-PIB uptake in Alzheimer's disease and MCI.

PURPOSE: The aim of this study was to evaluate the longitudinal changes in [(11)C]PIB uptake in mild cognitive impairment (MCI) and Alzheimer's disease (AD) over a long-term follow-up. METHODS: Six AD patients, ten MCI patients and eight healthy subjects underwent a [(11)C]PIB PET scan at baseline and at 2 and 5 years. The clinical status of the MCI patients was evaluated every 6 months. RESULTS: The MCI group showed a significant increase in [(11)C]PIB uptake over time (p < 0.001), with a similar increase from baseline to 2 years (4.7% per year) and from 2 to 5 years (5.0% per year). Eight MCI patients (80%) converted to AD, and two of these patients showed a normal [(11)C]PIB scan at baseline but increased uptake later. There was an increase in [(11)C]PIB uptake with time in the AD group (p = 0.02), but this did not significantly differ from the change in the control group. CONCLUSION: Our results revealed a significant increase in amyloid load even at the time of AD diagnosis in some of the MCI patients who converted. A positive [(11)C]PIB scan at baseline in MCI patients strongly predicted future conversion to AD but a negative PIB scan in MCI patients did not exclude future conversion. The results suggest that there is wide individual variation in the brain amyloid load in MCI, and in the course of amyloid accumulation in relation to the clinical diagnosis of AD.

[11C]PIB, [18F]FDG and MR imaging in patients with mild cognitive impairment.

PURPOSE: Cortical glucose metabolism, brain amyloid ß accumulation and hippocampal atrophy imaging have all been suggested as potential biomarkers in predicting which patients with mild cognitive impairment (MCI) will convert to Alzheimer's disease (AD). The aim of this study was to compare the prognostic ability of [(11)C]PIB PET, [(18)F]FDG PET and quantitative hippocampal volumes measured with MR imaging in predicting conversion to AD in patients with MCI. METHODS: The study group comprised 29 patients with MCI who underwent [(11)C]PIB PET and MR imaging. Of these, 22 also underwent [(18)F]FDG PET. All subjects were invited back for clinical evaluation after 2 years. RESULTS: During the follow-up time 17 patients had converted to AD while 12 continued to meet the criteria for MCI. The two groups did not differ in age, gender or education level, but the converter group tended to have lower MMSE and Word List learning than the nonconverter group. High [(11)C]PIB retention in the frontotemporal regions and anterior and posterior cingulate (p < 0.05) predicted conversion to AD. Also reduced [(18)F]FDG uptake in the left lateral temporal cortex (LTC) predicted conversion (p < 0.05), but quantitative hippocampal volumes did not (p > 0.1). In receiver operating characteristic (ROC) analysis the measurements that best predicted the conversion were [(11)C]PIB retention in the lateral frontal cortex and [(18)F]FDG uptake in the left LTC. Both PET methods resulted in good sensitivity and specificity and neither was significantly superior to the other. CONCLUSION: The findings indicate that [(11)C]PIB and [(18)F]FDG are superior to hippocampal volumes in predicting conversion to AD in patients with MCI.

The CMS19 disease model specifies a pivotal role for collagen XIII in bone homeostasis.

Mutations in the COL13A1 gene result in congenital myasthenic syndrome type 19 (CMS19), a disease of neuromuscular synapses and including various skeletal manifestations, particularly facial dysmorphisms. The phenotypic consequences in Col13a1 null mice (Col13a1-/-) recapitulate the muscle findings of the CMS19 patients. Collagen XIII (ColXIII) is exists as two forms, a transmembrane protein and a soluble molecule. While the Col13a1-/- mice have poorly formed neuromuscular junctions, the prevention of shedding of the ColXIII ectodomain in the Col13a1tm/tm mice results in acetylcholine receptor clusters of increased size and complexity. In view of the bone abnormalities in CMS19, we here studied the tubular and calvarial bone morphology of the Col13a1-/- mice. We discovered several craniofacial malformations, albeit less pronounced ones than in the human disease, and a reduction of cortical bone mass in aged mice. In the Col13a1tm/tm mice, where ColXIII is synthesized but the ectodomain shedding is prevented due to a mutation in a protease recognition sequence, the cortical bone mass decreased as well with age and the cephalometric analyses revealed significant craniofacial abnormalities but no clear phenotypical pattern. To conclude, our data indicates an intrinsic role for ColXIII, particularly the soluble form, in the upkeep of bone with aging and suggests the possibility of previously undiscovered bone pathologies in patients with CMS19.

PET amyloid ligand [11C]PIB uptake shows predominantly striatal increase in variant Alzheimer's disease.

Variant Alzheimer's disease (VarAD) with spastic paraparesis and presenile dementia is associated with certain mutations of the presenilin 1 (PS-1) gene, particularly those leading to deletion of exon 9 (PS-1Delta E9). VarAD is neuropathologically characterized by the presence of unusually large, Abeta42 positive, non-cored 'cotton wool' plaques (CWPs), also devoid of dystrophic neurites. The aim of the present study was to find out whether [(11)C]PIB would show increased uptake and serve as an in vivo biomarker of amyloid accumulation in VarAD. A further aim was to assess the correspondence of the [(11)C]PIB binding to the amount and type of Abeta deposits in another group of deceased VarAD patients' brains. We studied four patients with VarAD and eight healthy controls with PET using [(11)C]PIB as tracer. Parametric images were computed by calculating the region-to-cerebellum and region-to-pons ratio in each voxel over 60-90 min. Group differences in [(11)C]PIB uptake were analysed with automated region-of-interest (ROI) analysis. [(11)C]PIB uptake was compared to the immunohistochemically demonstrated deposition of Abeta in the brains of another group of four deceased VarAD patients. Patients with VarAD had significantly higher [(11)C] PIB uptake than the control group in the striatum (caudate nucleus and putamen), anterior and posterior cingulate gyrus, occipital cortex and thalamus. In the caudate and putamen [(11)C]PIB uptake, expressed as region-to-cerebellum ratio, was on the average 43% greater than the mean of the control group. The increases in the anterior (28%) and posterior (27%) cingulate gyrus, occipital cortex (21%) and thalamus (14%) were smaller. All VarAD patients showed this similar topographical pattern of increased [(11)C]PIB uptake. The results were essentially similar when the uptake was expressed as region-to-pons ratios. [(11)C]PIB imaging shows increased uptake in patients with VarAD especially in the striatum, and it can be used to detect amyloid accumulation in vivo in these patients. The pattern of increased [(11)C]PIB uptake is different from that described in sporadic Alzheimer's disease and resembles that seen in Alzheimer's disease patients with certain presenilin-1 mutations or amyloid precursor protein gene duplication showing predominantly striatal increase in [(11)C]PIB uptake.

Mutations in the LKB1 tumour suppressor are frequently detected in tumours from Caucasian but not Asian lung cancer patients.

Somatic mutations of LKB1 tumour suppressor gene have been detected in human cancers including non-small cell lung cancer (NSCLC). The relationship between LKB1 mutations and clinicopathological characteristics and other common oncogene mutations in NSCLC is inadequately described. In this study we evaluated tumour specimens from 310 patients with NSCLC including those with adenocarcinoma, adenosquamous carcinoma, and squamous cell carcinoma histologies. Tumours were obtained from patients of US (n=143) and Korean (n=167) origin and screened for LKB1, KRAS, BRAF, and EGFR mutations using RT-PCR-based SURVEYOR-WAVE method followed by Sanger sequencing. We detected mutations in the LKB1 gene in 34 tumours (11%). LKB1 mutation frequency was higher in NSCLC tumours of US origin (17%) compared with 5% in NSCLCs of Korean origin (P=0.001). They tended to occur more commonly in adenocarcinomas (13%) than in squamous cell carcinomas (5%) (P=0.066). LKB1 mutations associated with smoking history (P=0.007) and KRAS mutations (P=0.042) were almost mutually exclusive with EGFR mutations (P=0.002). The outcome of stages I and II NSCLC patients treated with surgery alone did not significantly differ based on LKB1 mutation status. Our study provides clinical and molecular characteristics of NSCLC, which harbour LKB1 mutations.

PET amyloid ligand [11C]PIB uptake and cerebrospinal fluid beta-amyloid in mild cognitive impairment.

BACKGROUND: In mild cognitive impairment (MCI), Alzheimer's disease (AD)-type cerebrospinal fluid (CSF) biomarker profiles predict rapid progression and conversion to AD. An increased brain amyloid burden in AD and MCI has been demonstrated with PET using [(11)C]PIB (Pittsburgh compound B). Little is known about the relationship between these biomarkers in MCI. METHODS: We studied 15 patients with amnestic MCI and 22 controls with PET using [(11)C]PIB. In MCI patients, CSF levels of Abeta42, pTAU, totalTAU and the Abeta42/pTAU ratio were measured. RESULTS: In MCI patients, CSF Abeta42 was abnormal in 53% of patients, totalTAU in 67%, pTAU in 64% and the Abeta42/pTAU ratio in 64%. A composite neocortical [(11)C]PIB uptake score was increased in 87% of the MCI patients. Only 54% of [(11)C]PIB-positive subjects showed AD-type Abeta42 values. During a 2-year follow-up, 6 MCI patients converted to AD, all of them had increased neocortical PIB scores at the MCI stage. Abnormal CSF Abeta42 was found in 3 patients, pTAU in 3 patients and Abeta42/pTAU ratio in 4 patients. CONCLUSION: Follow-up studies are needed to confirm whether [(11)C]PIB uptake might be more sensitive than CSF Abeta42 concentration in detecting increased amyloid burden in MCI, as suggested by the results of this study.

Dissolved air flotation (DAF) for primary and tertiary treatment of municipal wastewaters.

Tertiary treatment of municipal wastewater by dissolved air flotation was studied on a pilot-scale. The effects of coagulant dose, flocculation pattern, dispersion water recycle ratio and hydraulic surface load on process performance were evaluated. The treatment of primary effluents by dissolved air flotation was investigated to assess the suitability of this process for the treatment of heavily polluted effluents and wastewater treatment plant by-passes. The tertiary dissolved air flotation process typically achieved 90-99% reductions in the numbers of enteric microbes (total coliforms, enterococci and F-RNA coliphages). The average reductions of total phosphorus and chemical oxygen demand were 55-81% and 28-39%, respectively. Increasing the polyaluminium chloride coagulant dose from 2 to 10 mgAl(3+) l(-1) and the dispersion water recycle ratio from 11 to 22% improved the efficiency of the process. Changes in the flocculation conditions (range of G-values 10-55 s(-1); retention time 4-8 min) and hydraulic surface load (5 or 10 m h(-1)) did not clearly affect the process efficiency. The dissolved air flotation process decreased the numbers of enteric microbes and reduced total phosphorus, chemical oxygen demand and suspended solids from the primary treated wastewaters on average by 98-99.8%, 90%, 47% and 77%, respectively. The dissolved air flotation process was demonstrated to be a suitable method for efficient tertiary treatment of wastewaters, as well as for the elimination of peak pollution loads or by-pass wastewaters during the treatment plant overloading situations.

Peracetic acid (PAA) disinfection of primary, secondary and tertiary treated municipal wastewaters.

The efficiency of peracetic acid (PAA) disinfection against enteric bacteria and viruses in municipal wastewaters was studied in pilot-scale. Disinfection pilot-plant was fed with the primary or secondary effluent of Kuopio municipal wastewater treatment plant or tertiary effluent from the pilot-scale dissolved air flotation (DAF) unit. Disinfectant doses ranged from 2 to 7 mg/l PAA in the secondary and tertiary effluents, and from 5 to 15 mg/l PAA in the primary effluents. Disinfection contact times were 4-27 min. Disinfection of secondary and tertiary effluents with 2-7 mg/l PAA and 27 min contact time achieved around 3 log reductions of total coliforms (TC) and enterococci (EC). PAA disinfection also significantly improved the hygienic quality of the primary effluents: 10-15 mg/l PAA achieved 3-4 log reductions of TC and EC, 5 mg/l PAA resulting in below 2 log reductions. F-RNA coliphages were more resistant against the PAA disinfection and around 1 log reductions of these enteric viruses were typically achieved in the disinfection treatments of the primary, secondary and tertiary effluents. Most of the microbial reductions occurred during the first 4-18 min of contact time, depending on the PAA dose and microorganism. The PAA disinfection efficiency remained relatively constant in the secondary and tertiary effluents, despite of small changes of wastewater quality (COD, SS, turbidity, 253.7 nm transmittance) or temperature. The disinfection efficiency clearly decreased in the primary effluents with substantially higher microbial, organic matter and suspended solids concentrations. The results demonstrated that PAA could be a good alternative disinfection method for elimination of enteric microbes from different wastewaters.

Inactivation of enteric microorganisms with chemical disinfectants, UV irradiation and combined chemical/UV treatments.

The relative disinfection efficiencies of peracetic acid (PAA), hydrogen peroxide (H2O2) and sodium hypochlorite (NaOCl) against Escherichia coli, Enterococcus faecalis, Salmonella enteritidis and coliphage MS2 virus were studied in laboratory-scale experiments. This study also evaluated the efficiency of combined PAA/ultraviolet irradiation (UV) and H2O2/UV treatments to determine if the microbial inactivation was synergistic. Microbial cultures were added into a synthetic wastewater-like test medium and treated by chemical disinfectants with a 10 min contact time, UV irradiation or the combination of chemical and UV treatments. A peracetic acid dose of 3 mg/l resulted in approximately 2-3 log enteric bacterial reductions, whereas 7-15 mg/l PAA was needed to achieve 1-1.5 log coliphage MS2 reductions. Doses of 3-150 mg/l hydrogen peroxide achieved below 0.2 log microbial reductions. Sodium hypochlorite treatments caused 0.3-1 log microbial reductions at an 18 mg/l chlorine dose, while 2.6 log reductions of E. faecalis were achieved at a 12 mg/l chlorine dose. The results indicate that PAA could represent a good alternative to chlorine compounds in disinfection procedures, especially in wastewaters containing easily oxidizable organic matter. Hydrogen peroxide is not an efficient disinfectant against enteric microorganisms in wastewaters. The combined PAA/UV disinfection showed increased disinfection efficiency and synergistic benefits with all the enteric bacteria tested but lower synergies for the coliphage MS2. This suggests that this method could improve the efficiency and reliability of disinfection in wastewater treatment plants. The combined H2O2/UV disinfection only slightly influenced the microbial reductions compared to UV treatments and showed some antagonism and no synergies.

MEK and PI3K inhibition in solid tumors: rationale and evidence to date.

PI3K-AKT-mTOR and Ras-Raf-MEK-ERK are the most commonly altered oncogenic pathways in solid malignancies. There has been a lot of enthusiasm to develop inhibitors to these pathways for cancer therapy. Unfortunately, the antitumor activities of single-agent therapies have generally been disappointing, excluding B-Raf mutant melanoma and renal cell cancer. Preclinical studies have suggested that concurrent targeting of the PI3K-AKT-mTOR and Ras-Raf-MEK-ERK pathways is an active combination in various solid malignancies. In the current work, we review the preclinical data of the PI3K and MEK dual targeting as a cancer therapy and the results of early-phase clinical trials, and propose future directions.

New function for NF1 tumor suppressor.

The expression and subcellular localization of neurofibromatosis type 1 tumor suppressor was studied in keratinocytes induced to differentiate by increased Ca2+ concentration of the culture medium. Differentiating keratinocytes became intensely immunoreactive for neurofibromatosis type 1 protein, which was apparently associated with cellular fibrils. Double immunolabeling with antibodies to cytokeratin 14 and neurofibromatosis type 1 protein suggested an association of intermediate type cytoskeleton and neurofibromatosis type 1 protein. The presence of neurofibromatosis type 1 protein in cell preparations treated with cytoskeletal buffer indicated a high affinity interaction between intermediate filaments and neurofibromatosis type 1 protein. Further studies utilizing double immunolabelings revealed that the intense neurofibromatosis type 1 tumor suppressor signal on intermediate filaments was temporally limited to the period in keratinocyte differentiation in which the formation of desmosomes takes place. Keratinocytes were also cultured from nine patients with type 1 neurofibromatosis and were studied with respect to cell morphology, and association of neurofibromatosis type 1 protein with intermediate cytoskeleton. The results showed that keratinocytes cultured from patients with neurofibromatosis type 1 displayed a highly variable cell size and morphology compared to controls. The latter findings represent predicted alterations in a situation where cytoskeletal organization is disturbed. Furthermore, differentiating neurofibromatosis type 1 keratinocytes were characterized by a reduced number of cytokeratin bundles that were decorated neurofibromatosis type 1 protein. The results of this study suggest that neurofibromatosis type 1 tumor suppressor exerts its effects in part by controlling organization of cytoskeleton during the formation of cellular contacts.

Determination of salmonellae from municipal wastewaters.

This study compared the efficiency of culture methods for salmonellae detection in wastewaters collected from three Finnish municipal treatment plants and from one laboratory-scale plant. The performance of one-step enrichment in Preuss tetrathionate broth was better than that of two-step enrichment (buffered peptone water pre-enrichment (BPW) and selective enrichment in Rappaport-Vassiliadis (RV) medium. The best combinations for Salmonella isolation were xylose-lysine-deoxycholate (XLD) and Rambach (RB) agars after Preuss enrichment and did not differ when brilliant green-magnesium chloride (BM) or brilliant green phenol red (BP) agars were used. The two-step enrichment inhibited the growth of both salmonellae and interfering accompanying flora. Salmonella-positive plates were generally easier to read when inoculated from RV than from Preuss medium because of less growth of competing flora. XLD and BM agars supported growth of salmonellae and inhibited growth of competing flora better than BP and Rambach agars. XLD and BM agars gave the highest numbers of salmonellae isolations but XLD and Rambach agars gave the best differentiation. Salmonella levels were < 3- > 1100 MPN/100 mL.

Amyloid PET imaging in patients with mild cognitive impairment: a 2-year follow-up study.

BACKGROUND: Patients with amnestic mild cognitive impairment (MCI) have greater risk of conversion to Alzheimer disease (AD). Increased brain amyloid burden in AD and MCI has been demonstrated with PET using [(11)C] Pittsburgh compound B (PiB) as a tracer. OBJECTIVE: To evaluate change in ß-amyloid deposition in with MCI during 2-year follow-up. METHODS: Patients with MCI and controls were studied with [(11)C] PiB PET, MRI, and neuropsychometry at baseline and these investigations were repeated in patients with MCI after follow-up. RESULTS: Those patients with MCI converting to AD during follow-up had greater [(11)C] PiB retention in the posterior cingulate (p=0.020), in the lateral frontal cortex (p=0.006), in the temporal cortex (p=0.022), in the putamen (p=0.041), and in the caudate nucleus (p=0.025) as compared to nonconverters. In converters, there was no significant change in [(11)C] PiB uptake, whereas an increase was seen as compared to baseline in nonconverters in the anterior and posterior cingulate, temporal and parietal cortices, and putamen. Hippocampal atrophy was greater in converters at baseline than in nonconverters, but increased significantly in both groups during follow-up. CONCLUSIONS: Hippocampal atrophy and amyloid deposition seem to dissociate during the evolution of MCI, the atrophy increasing clearly and [(11)C] PiB retention changing modestly when conversion to AD occurs. Longer follow-up is needed to determine whether nonconverters would convert to AD later, which would suggest accelerated [(11)C] PiB retention preceding clinical conversion.

Conversion of amyloid positive and negative MCI to AD over 3 years: an 11C-PIB PET study.

BACKGROUND: Patients with amnestic mild cognitive impairment (MCI) represent an important clinical group as they are at increased risk of developing Alzheimer disease (AD). (11)C-PIB PET is an in vivo marker of brain amyloid load. OBJECTIVE: To assess the rates of conversion of MCI to AD during a 3-year follow-up period and to compare levels of amyloid deposition between MCI converters and nonconverters. METHODS: Thirty-one subjects with MCI with baseline (11)C-PIB PET, MRI, and neuropsychometry have been clinically followed up for 1 to 3 years (2.68 +/- 0.6 years). Raised cortical (11)C-PIB binding in subjects with MCI was detected with region of interest analysis and statistical parametric mapping. RESULTS: Seventeen of 31 (55%) subjects with MCI had increased (11)C-PIB retention at baseline and 14 of these 17 (82%) clinically converted to AD during follow-up. Only one of the 14 PIB-negative MCI cases converted to AD. Of the PIB-positive subjects with MCI, half (47%) converted to AD within 1 year of baseline PIB PET, these faster converters having higher tracer-retention values than slower converters in the anterior cingulate (p = 0.027) and frontal cortex (p = 0.031). Seven of 17 (41%) subjects with MCI with known APOE status were epsilon4 allele carriers, this genotype being associated with faster conversion rates in PIB-positive subjects with MCI (p = 0.035). CONCLUSIONS: PIB-positive subjects with mild cognitive impairment (MCI) are significantly more likely to convert to AD than PIB-negative patients, faster converters having higher PIB retention levels at baseline than slower converters. In vivo detection of amyloid deposition in MCI with PIB PET provides useful prognostic information.

Work-related symptoms and Salmonella antibodies among wastewater treatment plant workers.

Wastewater treatment plant workers are exposed to microbes, including Salmonella, but the prevalence of antibodies against Salmonella species or serovars in their serum samples has not been studied. Antibodies against Salmonella Infantis and lipopolysaccharide antigen common to S. Enteritidis and S. Typhimurium in immunoglobulin classes IgA, IgM and IgG were measured from 79 serum samples of wastewater treatment plant workers and from 79 blood donor samples. Faecal samples for Salmonella and Campylobacter were studied. Gastrointestinal, dermal and other symptoms were compared between 81 wastewater treatment plant workers and 89 food-processing workers. The blood donors had more antibodies against all of the tested antigens expect for S. Infantis in IgM and IgA classes, even though the wastewater treatment plant workers had more gastrointestinal symptoms than the controls. No Salmonella or Campylobacter were found in any faecal samples. Salmonella is not a probable cause of symptoms among wastewater treatment plant workers.

Keratinocytes cultured from patients with Hailey-Hailey disease and Darier disease display distinct patterns of calcium regulation.

BACKGROUND: Hailey-Hailey disease (HHD) (OMIM 16960) and Darier disease (DD) (OMIM 124200) are dominantly inherited acantholytic skin diseases, respectively, caused by mutations in the genes encoding the Golgi secretory pathway Ca2+-ATPase (SPCA1, ATP2C1) and the sarco/endoplasmic reticulum Ca2+-ATPase type 2 (SERCA2, ATP2A2) genes. OBJECTIVES: To investigate calcium regulation in keratinocytes cultured from patients with HHD and DD by measuring intracellular calcium resting levels and the cellular responses to ATP and thapsigargin. METHODS: The study was carried out using keratinocyte cultures established from four patients with HHD and four with DD. Calcium concentrations were measured with fluorescence ratio imaging using fura-2 loading. RESULTS: Control and HHD keratinocytes displayed approximately the same Ca2+ levels in resting phase, while DD keratinocytes showed elevated Ca2+ levels. Application of ATP caused less pronounced elevation of intracellular calcium concentration ([Ca2+]i) in both HHD and DD keratinocytes than in control cells. HHD keratinocytes did not lower their [Ca2+]i as efficiently as control keratinocytes after treatment with thapsigargin. In addition, DD keratinocytes were practically incapable of lowering their [Ca2+]i after treatment with thapsigargin. CONCLUSIONS: The results demonstrate that the defects in SPCA1 and SERCA2 calcium ATPases result in distinct patterns of calcium metabolism. This is also supported by the different clinical features of the diseases.

Neurofibromatosis type 1 tumour suppressor gene expression is deficient in psoriatic skin in vivo and in vitro: a potential link to increased Ras activity.

BACKGROUND: Neurofibromatosis type 1 (NF1) protein (neurofibromin) accelerates the inactivation of Ras-GTP in various cell types. Somatic mutations of the NF1 gene may lead to malignant transformation and uncontrolled proliferation. We have previously shown that NF1 protein expression is downregulated in psoriasis in vivo. OBJECTIVES: To study the functional expression and distribution of NF1 mRNA and protein in vivo and in psoriatic and normal keratinocyte cultures. METHODS: Immunohistochemistry and in situ hybridization were used to study NF1 gene and protein expression in psoriasis in vivo. Furthermore, Northern and in situ hybridizations, immunoblot and localization analyses were utilized to study NF1 mRNA and protein in vitro in keratinocyte cultures. RESULTS: NF1 tumour suppressor gene expression was reduced in lesional psoriatic skin compared with perilesional and normal skin in vivo. The in vitro results showed that the levels of NF1 mRNA and protein were reduced in cultured psoriatic keratinocytes during cellular differentiation even after multiple passaging of the cells. Moreover, cultured nonlesional psoriatic keratinocytes were almost equally defective as lesional cells with respect to NF1 expression. CONCLUSIONS: Our findings demonstrate that psoriatic keratinocytes maintain an altered phenotype and gene expression profile even when isolated from interaction with lymphocytes and fibroblasts, which are known to increase proliferation of keratinocytes. As NF1 protein is regarded as a Ras proto-oncogene regulator, the aberrant expression and distribution of NF1 protein and mRNA found in the present study may be causative to the previously described increased activation of Ras in psoriatic lesions, and relate to altered cellular behaviour.