Reducing the lipase LIPE in mutant α-synuclein mice improves Parkinson-like deficits and reveals sex differences in fatty acid metabolism.

Impaired lipid metabolism is a risk factor for Parkinson's disease (PD) and dementia with Lewy bodies (DLB) and can shift the physiological α-synuclein (αS) tetramer-monomer (T:M) ratio toward aggregation prone monomers. A resultant increase in phospho-serine 129+ αS monomers associating with excess mono- and polyunsaturated fatty acids contributes to the αS aggregation. We previously reported that decreasing the release of monounsaturated fatty acids (MUFAs) by reducing or inhibiting the hormone sensitive lipase (LIPE) reversed pathologic αS phosphorylation and improved soluble αS homeostasis in cultured αS triplication PD neurons and reduced DAergic neurodegeneration in a C.elegans αS model. However, assessing LIPE as a potential therapeutic target for progressive PD motor phenotypes has not been investigated. 3K αS mice, representing a biochemical and neuropathological amplification of the E46K fPD-causing mutation, have decreased αS T:M ratios, lipidic aggregates, and a L-DOPA responsive PD-like motor syndrome. Here, we reduced LIPE by crossings of 3K mice with LIPE null mice, which attenuated motor deficits in male LIPE+/- knockdown (LKD)-3K mice. Heterozygous LIPE reduction was associated with an improved αS T:M ratio, and dopaminergic neurotransmitter levels and fiber densities. In female 3K-LKD mice, an increase in pS129+ and larger lipid droplets (LDs) likely decreased the benefits seen in males. Reducing LIPE decreased MUFA release from neutral lipid storage, thereby reducing MUFA in phospholipid membranes with which αS interacts. Our study highlights fatty acid turnover as a therapeutic target for Lewy body diseases and support LIPE as a promising target in males. LIPE regulation represents a novel approach to mitigate PD and DLB risk and treat disease.

Trait-mediated responses of caterpillar communities to spongy moth outbreaks and subsequent tebufenozide treatments.

Outbreaks of the spongy moth Lymantria dispar can have devastating impacts on forest resources and ecosystems. Lepidoptera-specific insecticides, such as Bacillus thuringiensis var. kurstaki (BTK) and tebufenozide, are often deployed to prevent heavy defoliation of the forest canopy. While it has been suggested that using BTK poses less risk to non-target Lepidoptera than leaving an outbreak untreated, in situ testing of this assumption has been impeded by methodological challenges. The trade-offs between insecticide use and outbreaks have yet to be addressed for tebufenozide, which is believed to have stronger side effects than BTK. We investigated the short-term trade-offs between tebufenozide treatments and no-action strategies for the non-target herbivore community in forest canopies. Over 3 years, Lepidoptera and Symphyta larvae were sampled by canopy fogging in 48 oak stands in southeast Germany during and after a spongy moth outbreak. Half of the sites were treated with tebufenozide and changes in canopy cover were monitored. We contrasted the impacts of tebufenozide and defoliator outbreaks on the abundance, diversity, and functional structure of chewing herbivore communities. Tebufenozide treatments strongly reduced Lepidoptera up to 6 weeks after spraying. Populations gradually converged back to control levels after 2 years. Shelter-building species dominated caterpillar assemblages in treated plots in the post-spray weeks, while flight-dimorphic species were slow to recover and remained underrepresented in treated stands 2 years post-treatment. Spongy moth outbreaks had minor effects on leaf chewer communities. Summer Lepidoptera decreased only when severe defoliation occurred, whereas Symphyta declined 1 year after defoliation. Polyphagous species with only partial host plant overlap with the spongy moth were absent from heavily defoliated sites, suggesting greater sensitivity of generalists to defoliation-induced plant responses. These results demonstrate that both tebufenozide treatments and spongy moth outbreaks alter canopy herbivore communities. Tebufenozide had a stronger and longer lasting impact, but it was restricted to Lepidoptera, whereas the outbreak affected both Lepidoptera and Symphyta. These results are tied to the fact that only half of the outbreak sites experienced severe defoliation. This highlights the limited accuracy of current defoliation forecast methods, which are used as the basis for the decision to spray insecticides.

Survival of Norway spruce remains higher in mixed stands under a dryer and warmer climate.

Shifts in tree species distributions caused by climatic change are expected to cause severe losses in the economic value of European forestland. However, this projection disregards potential adaptation options such as tree species conversion, shorter production periods, or establishment of mixed species forests. The effect of tree species mixture has, as yet, not been quantitatively investigated for its potential to mitigate future increases in production risks. For the first time, we use survival time analysis to assess the effects of climate, species mixture and soil condition on survival probabilities for Norway spruce and European beech. Accelerated Failure Time (AFT) models based on an extensive dataset of almost 65,000 trees from the European Forest Damage Survey (FDS)--part of the European-wide Level I monitoring network--predicted a 24% decrease in survival probability for Norway spruce in pure stands at age 120 when unfavorable changes in climate conditions were assumed. Increasing species admixture greatly reduced the negative effects of unfavorable climate conditions, resulting in a decline in survival probabilities of only 7%. We conclude that future studies of forest management under climate change as well as forest policy measures need to take this, as yet unconsidered, strongly advantageous effect of tree species mixture into account.

Hybrid MCDA Methods to Integrate Multiple Ecosystem Services in Forest Management Planning: A Critical Review.

Multi-criteria decision analysis (MCDA) is a decision aid frequently used in the field of forest management planning. It includes the evaluation of multiple criteria such as the production of timber and non-timber forest products and tangible as well as intangible values of ecosystem services (ES). Hence, it is beneficial compared to those methods that take a purely financial perspective. Accordingly, MCDA methods are increasingly popular in the wide field of sustainability assessment. Hybrid approaches allow aggregating MCDA and, potentially, other decision-making techniques to make use of their individual benefits and leading to a more holistic view of the actual consequences that come with certain decisions. This review is providing a comprehensive overview of hybrid approaches that are used in forest management planning. Today, the scientific world is facing increasing challenges regarding the evaluation of ES and the trade-offs between them, for example between provisioning and regulating services. As the preferences of multiple stakeholders are essential to improve the decision process in multi-purpose forestry, participatory and hybrid approaches turn out to be of particular importance. Accordingly, hybrid methods show great potential for becoming most relevant in future decision making. Based on the review presented here, the development of models for the use in planning processes should focus on participatory modeling and the consideration of uncertainty regarding available information.

3 T MRI uterine peristalsis: comparison of symptomatic fibroid patients versus controls.

AIM: To compare uterine peristalsis between symptomatic fibroid patients and normal subjects and to determine the possible effect of fibroid characteristics on uterine peristalsis at high-field magnetic resonance imaging (MRI). MATERIALS AND METHODS: The present study included 20 symptomatic fibroid patients (age range 39-53 years) and 20 normal subjects (age range 19-46 years). MRI images were obtained during the peri-ovulatory phase using 3 T MRI using a sagittal T2 turbo spin-echo sequence and a half-Fourier acquisition single-shot turbo spin-echo sequence for display on cine mode. Two radiologists independently evaluated the images for the presence of uterine peristalsis by confidence level. In cases where peristalsis was present, the images were also evaluated for peristalsis frequency and direction. For fibroid patients, uterine and index fibroid volume, fibroid burden and index fibroid location were also recorded. RESULTS: Uterine peristalsis was significantly decreased in symptomatic fibroid patients compared with normal controls (p < 0.01). Peristalsis frequency in fibroid patients was also lower than in normal subjects. Direction of peristalsis was cervix-to-fundus for the majority of fibroid patients and controls. There was no significant relationship between fibroid characteristics, such as uterine volume, index fibroid volume, index fibroid location, and fibroid number in fibroid patients with, and fibroid patients without peristalsis. CONCLUSION: In women with symptomatic fibroids, the presence of uterine peristalsis is significantly decreased compared to normal controls on 3 T cine MRI. The presence of fibroids appears to disturb the normal conduction of uterine peristalsis and may interfere with fluid (e.g., menses, sperm) transport.

Efficacy of reciprocating and rotary NiTi instruments for retreatment of curved root canals assessed by micro-CT.

AIM: To compare the efficacy of reciprocating and rotary NiTi-instruments in removing filling material from curved root canals using micro-computed tomography. METHODOLOGY: Sixty curved root canals were prepared and filled with gutta-percha and sealer. After determination of root canal curvatures and radii in two directions as well as volumes of filling material, the teeth were assigned to three comparable groups (n = 20). Retreatment was performed using Reciproc, ProTaper Universal Retreatment or Hedström files. Percentages of residual filling material and dentine removal were assessed using micro-CT imaging. Working time and procedural errors were recorded. Statistical analysis was performed by variance procedures. RESULTS: No significant differences amongst the three retreatment techniques concerning residual filling material were detected (P > 0.05). Hedström files removed significantly more dentine than ProTaper Universal Retreatment (P < 0.05), but the difference concerning dentine removal between both NiTi systems was not significant (P > 0.05). Reciproc and ProTaper Universal Retreatment were significantly faster than Hedström files (P = 0.0001). No procedural errors such as instrument fracture, blockage, ledging or perforation were detected for Hedström files. Three perforations were recorded for ProTaper Universal Retreatment, and in both NiTi groups, one instrument fracture occured. CONCLUSIONS: Remnants of filling material were observed in all samples with no significant differences between the three techniques. Hedström files removed significantly more dentine than ProTaper Universal Retreatment, but no significant differences between both NiTi systems were detected. Procedural errors were observed with ProTaper Universal Retreatment and Reciproc.

Efficacy of D-RaCe and ProTaper Universal Retreatment NiTi instruments and hand files in removing gutta-percha from curved root canals - a micro-computed tomography study.

AIM: To compare the efficacy of two rotary NiTi retreatment systems and Hedström files in removing filling material from curved root canals. METHODOLOGY: Curved root canals of 57 extracted teeth were prepared using FlexMaster instruments and filled with gutta-percha and AH Plus. After determination of root canal curvatures and radii in two directions, the teeth were assigned to three identical groups (n = 19). The root fillings were removed with D-RaCe instruments, ProTaper Universal Retreatment instruments or Hedström files. Pre- and postoperative micro-CT imaging was used to assess the percentage of residual filling material as well as the amount of dentine removal. Working time and procedural errors were recorded. Data were analysed using analysis of covariance and analysis of variance procedures. RESULTS: D-RaCe instruments were significantly more effective than ProTaper Universal Retreatment instruments and Hedström files (P < 0.05). Hedström files removed significantly less dentine than the rotary NiTi systems (P < 0.0001). D-RaCe instruments were significantly faster compared to both other groups (P < 0.05). No procedural errors such as instrument fracture, blockage, ledging or perforation were detected in the Hedström group. In the ProTaper group, four instrument fractures and one lateral perforation were observed. Five instrument fractures were recorded for D-RaCe. CONCLUSIONS: D-RaCe instruments were associated with significantly less residual filling material than ProTaper Universal Retreatment instruments and hand files. Hedström files removed significantly less dentine than both rotary NiTi systems. Retreatment with rotary NiTi systems resulted in a high incidence of procedural errors.

Genes involved in senescence and immortalization.

Senescence is now understood to be the final phenotypic state adopted by a cell in response to several distinct cell physiological processes, including proliferation, oncogene activation and oxygen free radical toxicity. The role of telomere maintenance in immortalization and the roles of p16(INK4A), p19(ARF), p53 and other genes in senescence are being further elucidated. Significant progress continues to be made in our understanding of cellular senescence and immortalization.

Inhibition of telomerase limits the growth of human cancer cells.

Telomerase is a ribonucleoprotein enzyme that maintains the protective structures at the ends of eukaryotic chromosomes, called telomeres. In most human somatic cells, telomerase expression is repressed, and telomeres shorten progressively with each cell division. In contrast, most human tumors express telomerase, resulting in stabilized telomere length. These observations indicate that telomere maintenance is essential to the proliferation of tumor cells. We show here that expression of a mutant catalytic subunit of human telomerase results in complete inhibition of telomerase activity, reduction in telomere length and death of tumor cells. Moreover, expression of this mutant telomerase eliminated tumorigenicity in vivo. These observations demonstrate that disruption of telomere maintenance limits cellular lifespan in human cancer cells, thus validating human telomerase reverse transcriptase as an important target for the development of anti-neoplastic therapies.

Toll-like receptor 1 gene polymorphisms in childhood IgA nephropathy: a case-control study in the Korean population.

Toll-like receptors (TLRs) are innate immune mediators that stimulate nuclear factor kappa B and the inflammatory cytokines. TLR1 is expressed in renal tubular epithelial cells when the kidney is injured, but the role of TLR1 gene in glomerulonephritis has not been clearly elucidated. We aimed to investigate the association of TLR1 polymorphisms with immunoglobulin A nephropathy (IgAN) in children. One hundred and ninety pediatric patients with biopsy-proven IgAN and 283 healthy control subjects were enrolled. Two single nucleotide polymorphisms of TLR1 gene [rs4833095 (missense, Asn248Ser) and rs5743557 (promoter, -414C/T)] were selected and genotyped by direct sequencing. For rs4833095, the C/T genotype in the codominant model (vs. the T/T genotype) [odds ratio (OR) = 2.11, 95% confidence interval (CI): 1.21-3.69, P = 0.009] and the genotype containing C allele (C/T and C/C) in the dominant model (vs. the T/T genotype) (OR = 1.97, 95% CI: 1.16-3.34, P = 0.012) were associated with an increased risk of IgAN. For rs5743557, the T/T genotype in the codominant model (vs. the C/C genotype) (OR = 1.74, 95% CI: 1.02-2.96, P = 0.041) appeared to be associated with IgAN risk. In haplotype analysis, the CT haplotype revealed an association with IgAN (codominant model, OR = 1.38, 95% CI: 1.06-1.80, P = 0.017; dominant model, OR = 1.76, 95% CI: 1.16-2.67, P = 0.008). After Bonferroni correction, the association of the genotypes of rs4833095 and the CT haplotype with IgAN risk remained significant. These findings suggest that TLR1 gene polymorphisms may affect IgAN susceptibility in Korean children.

Naked DNA expressing two isoforms of hepatocyte growth factor induces collateral artery augmentation in a rabbit model of limb ischemia.

Hepatocyte growth factor (HGF) has been shown to induce angiogenesis in vivo and has potential as a candidate gene for 'therapeutic angiogenesis'. In vivo, two isoforms of HGF, HGF723 and HGF728, consisting of 723 and 728 amino acids, are generated through alternative splicing between exons 4 and 5, but the biological effects of their coexpression have not yet been elucidated. In this study, we generated a series of genomic-complementary DNA (cDNA) hybrids of the HGF gene by inserting various truncated intron 4 into the junction of exons 4 and 5 of HGF cDNA and analyzed the biological activities of these hybrid constructs. We showed that: (1) the hybrid called HGF-X7, which contained 1502 base pairs of intron 4, could drive a higher level of HGF expression than other hybrid constructs and cDNAs of each isoform alone; (2) the pCK vector was most efficient for the gene expression of HGF-X7; (3) coexpression of both isoforms of HGF could more efficiently induce the migration of human umbilical vein endothelial cell (HUVEC) and of the mouse myoblast cell line C2C12 myoblasts than a single isoform of HGF and human vascular endothelial growth factor (VEGF)165 at a given concentration; (4) intramuscular administration of pCK-HGF-X7 resulted in transient and localized HGF expression in the injected muscle without an increase in the HGF protein levels in other tissues including serum; and (5) intramuscular injection of pCK-HGF-X7 could more efficiently increase the number of angiographically recognizable collateral vessels, as well as improve an intra-arterial Doppler wire-measured blood flow in the rabbit model of hindlimb ischemia when compared with the identical vector encoding VEGF165 gene. These results showed that transfer of the genomic-cDNA hybrid of the HGF gene could be used as a potential therapeutic approach to human vascular diseases.

Separable portions of the CD2 cytoplasmic domain involved in signaling and ligand avidity regulation.

Effective T cell immune responses require the molecular interplay between adhesive and signaling events mediated by the T cell receptor for antigen (TCR) and other cell surface coreceptor molecules. In this report, we have distinguished between the role of regulated adhesion and transmembrane signaling in coreceptor function using the T cell glycoprotein CD2. By binding its ligands on antigen-presenting cell (APC), CD2 serves both to initiate signal transduction events and to promote cellular adhesion. Furthermore, the avidity of CD2 for one ligand, CD58 (LFA-3), is regulated by TCR signaling. We have expressed wild type CD2 and a series of mutated CD2 molecules in an antigen-specific murine T cell hybridoma. Structure-function studies using these stably transfected cell lines identify two structurally and functionally distinct regions of the 116 amino acid (aa) cytoplasmic domain. One region is required for CD2-mediated signal transduction, and a separate COOH-terminal 21 aa portion is required for CD2 activity regulation. Cell lines expressing CD2 molecules lacking the cytoplasmic segment required for CD2-initiated IL-2 production retain the ability to upregulate CD2 avidity. Conversely, cell lines expressing CD2 mutants lacking the cytoplasmic segment required for avidity regulation retain the ability to initiate CD2-specific signaling. In antigen-specific T cell responses, basal binding of CD2 to its ligands enhances antigen responsiveness only minimally, whereas regulated avidity and transmembrane signaling are both required for optimal coreceptor function. Taken together, these studies demonstrate the independent contributions of regulated adhesion and intracellular signaling in CD2 coreceptor function.

Mesosecrin: a secreted glycoprotein produced in abundance by human mesothelial, endothelial, and kidney epithelial cells in culture.

Human mesothelial cells, endothelial cells, and type II kidney epithelial cells growing in culture devote approximately 3% of their total protein synthesis to the production of an Mr approximately 46-kD, pI 7.1, secreted glycoprotein (designated Sp46). Fibroblasts make about 1/10th as much Sp46 as these cell types, and their synthesis is dependent upon hydrocortisone. Keratinocytes, urothelial cells, conjunctival epithelial cells, and mammary epithelial cells do not make detectable amounts of Sp46. Mesothelial cells secrete Sp46 onto the substratum, and from there it is subsequently released into the medium. Immunofluorescence analysis using specific antisera discloses that Sp46 is deposited beneath cells as a fine coating on the substratum. In sparse cultures, Sp46 is detected in trails behind motile cells. In contrast, secreted fibronectin coalesces into fibers, most of which remain in contact with and on top of the cells; thus Sp46 does not preferentially bind to fibronectin. About 6 kD of the mass of human Sp46 is N-linked oligosaccharide, which is terminally sialated before secretion. Sp46 has a low glycine content, indicating that it is not a collagenlike protein. Its NH2-terminal sequence over the first 40 amino acids does not resemble any protein for which sequence information is available. Sp46 appears to be a novel extracellular glycoprotein, high-level constitutive expression of which is restricted to mesoderm-derived epithelial and endothelial cells. We therefore propose for it the name "mesosecrin."

Genetic expression in the developing brain.

The adult mouse brain contains complex populations of polyadenylated [poly(A)+] and nonpolyadenylated [poly(A)-] messenger RNA's (mRNA's). These mRNA's are separate sequence populations, similar in complexity, and in combination are equivalent to approximately 150,000 different mRNA sequences, of average length. Essentially all of the "adult" poly(A)+ mRNA's are present in the brain at birth. In contrast, most of the poly(A)- mRNA's are absent. Brain poly(A)- mRNA's begin to appear soon after birth, but the full adult complement is not reached until young adulthood. This suggests that these poly(A)- mRNA's specify proteins required for the biological capabilities of the brain that emerge during the course of postnatal development.

Brain "identifier sequence" is not restricted to brain: similar abundance in nuclear RNA of other organs.

A repeated 82 base pair sequence in genomic DNA of the rat was previously proposed as being a control element governing brain (neuron) specific genetic expression. This intronic sequence, termed the brain "identifier" (ID), is complementary to small RNA species localized in brain cytoplasm, and it was thought to be represented specifically in RNA produced by brain nuclei in vitro. The RNA blot analyses of total nuclear and polyadenylated heterogeneous nuclear RNA described in the present report show that this ID sequence is also present in the liver and kidney in abundances similar to those in the brain. This repeated sequence is not, therefore, restricted to transcripts produced in the brain as suggested from previous transcriptional "runoff" experiments. Measurements on rat and mouse nuclear RNA indicate that the abundance of ID sequence transcript is roughly proportional to the number of copies of this repeat in the respective genomes. This suggests a rather random genomic location and transcription of this sequence. From these results it seems improbable that the ID sequence functions as a transcriptional-level control element in genes expressed specifically in the brain.

Transcription of nonrepeated DNA in mouse brain.

Under normal conditions of DNA renaturation, about 60 percent of mouse DNA fragments renature at a rate consistent with their being present only once per sperm. These nonrepeated sequences (also called single-copy or unique) may be used in RNA-DNA hybridization experiments to provide quantitative estimates of RNA diversity. About 10 percent of the mouse single-copy sequences are transcribed in mouse brain tissue. Estimates of about 3 percent were obtained for mouse liver and kidney RNA's. If only one of the complementary DNA strands is transcribed, this hybridization value implies that the equivalent of at least 300,000 different sequences of 1000 nucleotides are expressed in mouse brain tissue. It is suggested that the large amount of DNA in mammals is functionally important, and that a substantial proportion of the genome is expressed in the brain.

One strand equivalent of the Escherichia coli genome is transcribed: complexity and abundance classes of mRNA.

DNA-RNA hybridization experiments show that essentially all of the genomic information is transcribed. High, intermediate, and rare abundance classes of messenger RNA (mRNA) are present, and their estimated complexities are equal to about 240, 1300, and 700 average-sized mRNA species, respectively. The high abundance mRNA species are present, an average, two to three copies per cell and constitute about 95 percent of the mRNA mass. Intermediate abundance mRNA species are present, on average, about once per 35 cells. The relative abundance and complexity of these mRNA classes correspond well with previous respective measurements on protein. Rare RNA species are thought to represent maximally repressed genes. Analysis of RNA synthesized in vitro by isolated nucleoids (chromosomes) suggests that sense and nonsense sequences are extensively interspersed on a given strand of the DNA.

Overlapping but nonidentical binding sites on CD2 for CD58 and a second ligand CD59.

The interaction of the T cell glycoprotein CD2 with one ligand, CD58, contributes to T cell function. We have identified CD59, a glycoprotein with complement-inhibitory function, as a second physiological ligand for CD2. Antibodies to CD59 inhibit CD2-dependent T cell activation in murine T cell hybridomas expressing human CD2. In an in vitro binding assay with purified CD58 and CD59, CD2+ cells bind not only immobilized CD58 but also CD59. With two complementary approaches, it was demonstrated that the binding sites on CD2 for CD58 and CD59 are overlapping but nonidentical. These observations suggest that direct interactions between CD2 and both CD58 and CD59 contribute to T cell activation and adhesion.

The 46{degrees} Halo and Its Arcs.

Ice crystals in the form of right hexagonal prisms have faces that form 90 degrees prisms. Light rays were traced through these prism faces by computer calculation, and the light patterns that would be produced in the sky for a particular distribution of crystal orientations were simulated. Crystals with random orientations produce a 46 degrees halo. Hexagonal plate crystals with nearly horizontal end faces produce circumzenithal and circumhorizontal arcs. Hexagonal column crystals with horizontal axes produce supralateral and infralateral arcs. Plate crystals spinning about a horizontal axis that is a face diagonal of the crystal produce a series of arcs touching the 46 degrees halo. Each of these effects was simulated for several elevations of the sun.

Cyclin D1 overexpression and p53 inactivation immortalize primary oral keratinocytes by a telomerase-independent mechanism.

The immortalization of human cells is a critical step in multistep carcinogenesis. Oral-esophageal carcinomas, a model system to investigate molecular mechanisms underlying squamous carcinogenesis, frequently involve cyclin D1 overexpression and inactivation of the p53 tumor suppressor. Therefore, our goal was to establish the functional role of cyclin D1 overexpression and p53 inactivation in the immortalization of primary human oral squamous epithelial cells (keratinocytes) as an important step toward malignant transformation. Cyclin D1 overexpression alone was found to induce extension of the replicative life span of normal oral keratinocytes, whereas the combination of cyclin D1 overexpression and p53 inactivation led to their immortalization. This study also demonstrates that immortalization of oral keratinocytes can be independent of telomerase activation, involving an alternative pathway of telomere maintenance (ALT).