Die Rolle der Autophagie im Knochenremodellierung

Background and Aims: Accumulating evidence suggests a key-role for autophagy in various diseases via regulation of cellular metabolism and differentiation. However, the functional role of autophagy in the pathogenesis of bone- and joint diseases is not well understood. The first aim of this thesis was to investigate the role of autophagy in two bone diseases with pathologically increased generation of bone-degrading osteoclasts, rheumatoid arthritis and osteoporosis. We analyzed the role of autophagy in osteoclast differentiation and activity as well as the therapeutic effects of targeting autophagy. The second aim of our study is to characterize the role of autophagy in osteoarthritis, a joint disease with progressive loss of articular chondrocytes and proteoglycans and changes of the underlying bone. We characterized the outcome of mice with targeted inactivation of autophagy in chondrocytes in an experimental model of osteoarthritis. Methods: Atg7fl/fl x LysMCre and Atg7fl/fl x Col2a1Cre mice were generated to specifically inhibit autophagy in osteoclasts and chondrocytes, respectively. Autophagy was also inhibited pharmacologically by treatment with chloroquin (CQ). hTNF tg (human tumor necrosis factor alpha transgenic) mice, which resemble characteristic findings of human rheumatoid arthritis, served as a model of inflammatory bone loss. Non-inflammatory bone loss was induced by ovariectomy (OVX). Surgical destabilization of the medial meniscus (DMM) was performed in mice to mimic human osteoarthritis. The outcomes were analysed by microcomputed tomography (μCT), histomorphometry, Tartrate-resistant acid phosphatase staining (TRAP, to identify osteoclasts), safranin O and Toluidine blue stainings, rt-qPCR and immunohistochemistry for osteoclasts and chondrocytes and autophagy associated genes and markers. Results: We first demonstrated that autophagy is activated in osteoclasts of RA patients and that this activation might be mediated by TNFα. Pharmacologic or genetic inactivation of autophagy downregulated the expression of osteoclast markers such as NFATc1, OSCAR and prevented osteoclast differentiation in vitro and in vivo. hTNFα tg mice transplanted with Atg7fl/fl x LysMCre+ bone marrow were protected from TNFα induced local and systemic bone loss. Moreover, knockdown of Atg7 in osteoclast precursors or treatment with CQ also prevented OVX bone loss with reduced osteoclast numbers and maintained the bone density. Abstract 2 In the other part of the study, we observed that the expression of autophagy related genes, Atg7 and Beclin1, are down regulated in articular chondrocytes in experimental osteoarthritis. Targeted deletion of Atg7 in chondrocytes accelerated the loss of proteoglycans and promotes osteophyte formation in the DMM model. Selective inhibition of autophagy in chondrocytes leads to increased accumulation of type X collagen, providing evidence that autophagy regulations hypertrophic differentiation of chondrocytes. Conclusion: We demonstrated that autophagy is involved in the pathogenesis of rheumatoid arthritis, osteoporosis and osteoarthritis. Autophagy is essential for osteoclast differentiation and bone resorption activity. Inhibition of autophagy effectively inhibited osteoclastogenesis and ameliorated local and systemic bone loss in murine models of RA and osteoporosis. Autophagy is also an important regulator of chondrocyte homeostasis and inactivation of autophagy in chondrocytes accelerates experimental osteoarthritis Considering that inhibitors of autophagy such as CQ approved for clinical use, these findings may have direct translational implications and may point to novel strategies for the treatment of these bone and joint diseases.Hintergrund und Ziele: Autophagie spielt in vielen Erkrankungen eine Schlüsselrolle, da sie Zelldifferenzierung- und Überleben reguliert. Die Bedeutung der Autophagie in der Entstehung von Knochenerkrankungen ist jedoch weitestgehend unbekannt. Im ersten Teil meiner Arbeit untersuchte ich daher die Rolle der Autophagie in der rheumatoiden Arthritis und der Osteoporose, zweier kataboler Knochenerkrankungen, in denen das Gleichgewicht von Knochenabbau durch Osteoklasten und Knochenaufbau durch Osteoblasten wesentlich gestört ist. Ziele meiner Untersuchungen sind es daher die Bedeutung der Autophagie für Osteoklastendifferenzierung- und Aktivität besser zu verstehen und die Modulation der Autophagie als möglichen Therapieansatz zur Behandlung der rheumatoiden Arthritis und der Osteoporose in präklinischen Modellen zu analysieren. Im zweiten Teil zielen wir eine bessere Verständnis in der Pathogenese von Osteoarthritis, eine Knochenerkrankung charakterisiert mit progressiven artikuläre Chondrozyten und Proteoglykaneverlust und Veränderungen der drunter liegende Knochen GewebeDazu benützen wir ein Experimentelle Maus Modelle der die Humane Osteoarthritis nachahmt. Methoden: Atg7fl/fl x LysMCre und Atg7fl/fl x Col2a1Cre Mäuse wurden generiert um Autophagie spezifisch in Osteoklasten und Chondrozyten auszuschalten. Neben diesem genetischen Ansatz, wurde Autophagie auch pharmakologisch mit Chloroquin (CQ) gehemmt. Mäuse transgen für humanes TNFα (tumor necrosis factor alpha), die einen Phänotyp entwickeln, der der humanen rheumatoiden Arthritis ähnelt, wurden als Modell für inflammatorischen Knochenabbau genutzt. Das Ovarektomiemodell (OVX) repräsentierte einen nicht-inflammatorische Knochenverlust, wie er bei der postmenopausale Osteoporose auftritt. Chirurgische Destabilisierung des medialen Meniskus (DMM) wurde als Modell für die Osteoarthrose genutzt. Analysiert wurden die Proben mit Mikro-Computertomografie (μCT), Histomorphologie, sowie folgenden immunhistochemischen Färbungen: Tartrateresistant acid phosphatase (TRAP), Safranin O, Toluidin-Blau. Daneben wurden real-time PCR (qPCR)-Messungen durchgeführt, um die Osteoklasten-, Chondozyten- und Autophagieassoziierten Gene zu bestimmen. Ergebnisse: Ich konnte zeigen, dass Autophagie in Osteoklasten von Patienten mit rheumatoider Arthritis TNFα-abhängig aktiviert ist. Sowohl pharmakologische als auch genetische Inaktivierung von Autophagie reduzierten die Expression von Osteoklastenmarkern, wie NFATc1 und OSCAR, und verhinderten somit die Osteoklastendifferenzierung in vitro und in vivo. hTNFα transgene Mäuse transplantiert mit Atg7fl/fl x LysMCre+ Knochenmark Zellen waren vor lokalem und systemischem Knochenabbau geschützt. Knockdown von Atg7 in monozytären Zellen und die Behandlung mit CQ konnten effektiv OVX-induzierten Knochenverlust verhindern. μCT-Analysen zeigten, dass Hemmung der Autophagie Knochenverlust in der DMM Modell entgegenwirkt. Die Expression der Autophagie-assozierte Gene Atg7 und Beclin1 in artikulären Chondrozyten war im im DMM-Modell vermindert. Spezifische Deletion von Atg7 in Chondrozyten verstärkt den Knorpel- und Matrixverlust sowie die Osteophytenbildung in diesem Modell und führte zu einer vermehrten hypertropher Differenzierung der Chondrozyten. Schlussfolgerung: Diese Ergebnisse zeigen eine Vielzahl verschiedener Funktionen der Autophagie in verschieden Knochenerkrankungen aufdecken. Meine Ergebnisse zeigen, dass Autophagie eine große Rolle in der Differenzierung von Osteoklasten und daraus resultierend in Knochenabbau-Prozessen spielt. Diese neugewonnen Kenntnisse ließen sich therapeutisch nutzen: Hemmung der Autophagie reduziert die Osteoklastogenese und den Knochenverlust in Modellen für RA und Osteoporose. Neben der Bedeutung in den Osteoklasten, hat die Autophagie eine Schlüsselrolle in der Chondrozytenhomöostase inne. Hemmung der Autophagie in Chondrozyten beschleunigt die Osteoarthrose

Implications of regional surface ozone increases on visibility degradation in southeast China

Long-term visibility (1968–2010) and air pollutant (1984–2010) data records in Hong Kong reveal that the occurrence of reduced visibility (RV, defined as the percentage of hours per month with visibility below 8 km in the absence of rain, fog, mist or relative humidity above 95%) in southeast China has increased significantly in the last four decades. The most pronounced rate of increase was observed after 1990 (nine times higher than that before 1990), when notable increases in surface ozone (O3) levels were simultaneously observed (1.06 µg m−3 per yr). The greatest increases in RV, and in O3, NO2 and SO2 concentrations are coincident in the autumn (1.47, 0.20 and 0.45 µg m−3 per yr respectively), when southeast China is strongly influenced by regional O3 formation and accumulation due to continental outflow of pollution from the east China coast under favourable meteorological conditions. Multiple regression revealed that the RV percentage correlated well (p<0.05) with NO2 and NO x in the 1980s, and with NO2, SO2 and O3 after the 1990s, suggesting that there have been changes in the predominant factors causing visibility degradation. In order to elucidate the reasons for these changes, the results were integrated with data from previous research. Possible impacts of elevated O3 on secondary particle formation and their effects on visibility degradation and aerosol radiative forcing in an oxidant-enhanced southeast China are highlighted. Other factors potentially leading to visibility degradation, such as ship emissions and biomass burning, are also discussed

Molecular population genetics and gene expression analysis of duplicated CBF genes of Arabidopsis thaliana

<p>Abstract</p> <p>Background</p> <p><it>CBF/DREB </it>duplicate genes are widely distributed in higher plants and encode transcriptional factors, or CBFs, which bind a DNA regulatory element and impart responsiveness to low temperatures and dehydration.</p> <p>Results</p> <p>We explored patterns of genetic variations of <it>CBF1, -2</it>, and -<it>3 </it>from 34 accessions of <it>Arabidopsis thaliana</it>. Molecular population genetic analyses of these genes indicated that <it>CBF2 </it>has much reduced nucleotide diversity in the transcriptional unit and promoter, suggesting that <it>CBF2 </it>has been subjected to a recent adaptive sweep, which agrees with reports of a regulatory protein of <it>CBF2</it>. Investigating the ratios of K_a/K_sbetween all paired <it>CBF </it>paralogus genes, high conservation of the AP2 domain was observed, and the major divergence of proteins was the result of relaxation in two regions within the transcriptional activation domain which was under positive selection after <it>CBF </it>duplication. With respect to the level of <it>CBF </it>gene expression, several mutated nucleotides in the promoters of <it>CBF3 </it>and <it>-1 </it>of specific ecotypes might be responsible for its consistently low expression.</p> <p>Conclusion</p> <p>We concluded from our data that important evolutionary changes in <it>CBF1, -2</it>, and -<it>3 </it>may have primarily occurred at the level of gene regulation as well as in protein function.</p

Comorbidity and dementia: A nationwide survey in Taiwan

Background Comorbid medical diseases are highly prevalent in the geriatric population, imposing hardship on healthcare services for demented individuals. Dementia also complicates clinical care for other co-existing medical conditions. This study investigated the comorbidities associated with dementia in the elderly population aged 65 years and over in Taiwan. Methods We conducted a nationwide, population-based, cross-sectional survey; participants were selected by computerized random sampling from all 19 Taiwan counties between December 2011 and March 2013. After exclusion of incomplete or erroneous data, 8,456 subjects were enrolled. Of them, 6,183 were cognitively normal (control group), 1,576 had mild cognitive impairment (MCI), and 697 had dementia. We collected information about types of comorbidities (i.e., vascular risk factors, lung diseases, liver diseases, gastrointestinal diseases, and cancers), Charlson comorbidity index score, and demographic variables to compare subjects with normal cognition, MCI, and dementia. Results Regardless of the cognitive condition, over 60% of the individuals in each group had at least one comorbid disease. The proportion of subjects possessing at least three comorbidities was higher in those with cognitive impairment (MCI 20.9%, dementia 27.3%) than in control group (15%). Hypertension and diabetes mellitus were the most common comorbidities. The mean number of comorbidities and Charlson comorbidity index score were greater in MCI and dementia groups than in control group. Logistic regression demonstrated that the comorbidities significantly associated with MCI and dementia were cerebrovascular disease (OR 3.35, CI 2.62–4.28), cirrhosis (OR 3.29, CI 1.29–8.41), asthma (OR 1.56, CI 1.07–2.27), and diabetes mellitus (OR 1.24, CI 1.07–1.44). Conclusion Multiple medical comorbid diseases are common in older adults, especially in those with cognitive impairment. Cerebrovascular disease, cirrhosis, asthma, and diabetes mellitus are important contributors to cognitive deterioration in the elderly. Efforts to lower cumulative medical burden in the geriatric population may benefit cognitive function

Comorbidity and dementia: A nationwide survey in Taiwan

Background Comorbid medical diseases are highly prevalent in the geriatric population, imposing hardship on healthcare services for demented individuals. Dementia also complicates clinical care for other co-existing medical conditions. This study investigated the comorbidities associated with dementia in the elderly population aged 65 years and over in Taiwan. Methods We conducted a nationwide, population-based, cross-sectional survey; participants were selected by computerized random sampling from all 19 Taiwan counties between December 2011 and March 2013. After exclusion of incomplete or erroneous data, 8,456 subjects were enrolled. Of them, 6,183 were cognitively normal (control group), 1,576 had mild cognitive impairment (MCI), and 697 had dementia. We collected information about types of comorbidities (i.e., vascular risk factors, lung diseases, liver diseases, gastrointestinal diseases, and cancers), Charlson comorbidity index score, and demographic variables to compare subjects with normal cognition, MCI, and dementia. Results Regardless of the cognitive condition, over 60% of the individuals in each group had at least one comorbid disease. The proportion of subjects possessing at least three comorbidities was higher in those with cognitive impairment (MCI 20.9%, dementia 27.3%) than in control group (15%). Hypertension and diabetes mellitus were the most common comorbidities. The mean number of comorbidities and Charlson comorbidity index score were greater in MCI and dementia groups than in control group. Logistic regression demonstrated that the comorbidities significantly associated with MCI and dementia were cerebrovascular disease (OR 3.35, CI 2.62–4.28), cirrhosis (OR 3.29, CI 1.29–8.41), asthma (OR 1.56, CI 1.07–2.27), and diabetes mellitus (OR 1.24, CI 1.07–1.44). Conclusion Multiple medical comorbid diseases are common in older adults, especially in those with cognitive impairment. Cerebrovascular disease, cirrhosis, asthma, and diabetes mellitus are important contributors to cognitive deterioration in the elderly. Efforts to lower cumulative medical burden in the geriatric population may benefit cognitive function

Differential evolution with two-level parameter adaptation

The performance of differential evolution (DE) largely depends on its mutation strategy and control parameters. In this paper, we propose an adaptive DE (ADE) algorithm with a new mutation strategy DE/lbest/1 and a two-level adaptive parameter control scheme. The DE/lbest/1 strategy is a variant of the greedy DE/best/1 strategy. However, the population is mutated under the guide of multiple locally best individuals in DE/lbest/1 instead of one globally best individual in DE/best/1. This strategy is beneficial to the balance between fast convergence and population diversity. The two-level adaptive parameter control scheme is implemented mainly in two steps. In the first step, the population-level parameters F p and CR p for the whole population are adaptively controlled according to the optimization states, namely, the exploration state and the exploitation state in each generation. These optimization states are estimated by measuring the population distribution. Then, the individual-level parameters F i and CR i for each individual are generated by adjusting the population-level parameters. The adjustment is based on considering the individual's fitness value and its distance from the globally best individual. This way, the parameters can be adapted to not only the overall state of the population but also the characteristics of different individuals. The performance of the proposed ADE is evaluated on a suite of benchmark functions. Experimental results show that ADE generally outperforms four state-of-the-art DE variants on different kinds of optimization problems. The effects of ADE components, parameter properties of ADE, search behavior of ADE, and parameter sensitivity of ADE are also studied. Finally, we investigate the capability of ADE for solving three real-world optimization problems

Correction of Tetralogy of Fallot with Absent Pulmonary Valve Syndrome in a Young Infant Using a Bicuspid Equine Pericardial Tube

Absent pulmonary valve syndrome (APVS) is an uncommon variant of tetralogy of Fallot (TOF), which manifests morphologically as vestigial pulmonary valve cusps at the right ventricle-pulmonary trunk junction. The aneurysmally dilated pulmonary arteries may compress the tracheobronchial tree and cause severe respiratory distress in the neonatal or infant stage. Early surgical correction in these patients is necessary despite the high operative mortality rate. A 1-day-old male neonate suffered from progressive shortness of breath after birth. Echocardiography confirmed the diagnosis of TOF with APVS. The marked dilatation of pulmonary arteries resulted in airway compression in addition to heart failure. Total surgical correction was performed at 40 days of age, using a homemade bicuspid equine pericardial tube for right ventricular outflow reconstruction. The short-term follow-up echocardiogram demonstrated good motility of the pericardial leaflet. However, patients receiving this type of valved conduit require meticulous long-term follow-up

Electronic Structures and Optical Properties of Phenyl C 71

Phenyl C71 butyric acid methyl ester (PC71BM) has been adopted as electron acceptor materials in bulk heterojunction solar cells with relatively higher power conversion efficiency. The understanding of the mechanism and performance for the devices based upon PC71BM requires the information of conformations, electronic structures, optical properties, and so forth. Here, the geometries, IR and Raman, electronic structures, polarizabilities, and hyperpolarizabilities of PC71BM isomers are studied by using density functional theory (DFT); the absorption and excitation properties are investigated via time-dependent DFT with B3LYP, PBE0, and CAM-B3LYP functionals. The calculated results show that [6,6]PC71BM is more stable than [5,6]PC71BM due to the lower total energy. The vibrational modes of the isomers at IR and Raman peaks are quite similar. As to absorption properties, CAM-B3LYP functional is the suitable functional for describing the excitations of PC71BM because the calculated results with CAM-B3LYP functional agree well with that of the experiment. The analysis of transition configurations and molecular orbitals demonstrated that the transitions at the absorption maxima in UV/Vis region are localized π-π* transitions in fullerenes cages. Furthermore, the larger isotropic polarizability of PC71BM indicates that the response of PC71BM to applied external electric field is stronger than that of PC61BM, and therefore resulting into better nonlinear optical properties