Optimierung von Transformationsprotokollen und Entwicklung von SSR-Markern für das Kolumnarwachstum beim Apfel

Ziel war es, molekularbiologische Untersuchungen zum Kolumnarwachstum des Apfels durchzuführen. Anhand Sequenzdaten des ‘Golden Delicious’ Genoms (Velasco et al. 2010) wurden drei neue SSR Marker entwickelt. Sie konnten bei untersuchten Geisenheimer Nachkommenschaften zuverlässig den Kolumnarwuchs auf DNA-Ebene detektieren. Zusätzlich wurden von Bai et al. (2012) veröffentlichte Marker untersucht. Die von Bai et al. (2012) gefundenen Grenzen des co-Lokus konnten in dieser Arbeit anhand der Geisenheimer Nachkommenschaften nicht bestätigt werden. Die „linke“ Begrenzung der co-Region wird nach Untersuchungen dieser Arbeit am ehesten von dem Marker Mdo.chr10.11 (Moriya et al. 2012) bei 18,757 Mbp definiert. Die „rechte“ Begrenzung der co-Region wird vermutlich von den Markern Co04R13 (Baldi et al. 2012) und C1753-3520 (Bai et al. 2012) bei 18,905 Mbp definiert, wodurch die potentielle co-Region auf 148 kb auf Chromosom 10 eingegrenzt werden könnte. Für Funktionsanalysen möglicher Kandidatengene des co-Gens wurde ein Agrobakterien-vermitteltes Transformationssystem für die Geisenheimer Apfelselektionen ‘A 14’ und ‘Procats 28’ adaptiert. Zusätzlich wurde der bereits in der Literatur als transformierbar beschriebene Genotyp ‘Jonagold’ (Viss et al. 2003) transformiert. Bei Transformationen der Apfelselektion ‘A 14’ gelang es, transgene Zellen an den Explantaten, am Kallusgewebe und an den Regeneraten zu erzeugen. Bei Transformationen von ‘Jonagold’ wurde ein fast vollständig transgenes Regenerat erzeugt.

Debris-flow activity in abandoned channels of the Manival torrent reconstructed with LiDAR and tree-ring data

Hydrogeomorphic processes are a major threat in many parts of the Alps, where they periodically damage infrastructure, disrupt transportation corridors or even cause loss of life. Nonetheless, past torrential activity and the analysis of areas affected during particular events remain often imprecise. It was therefore the purpose of this study to reconstruct spatio-temporal patterns of past debris-flow activity in abandoned channels on the forested cone of the Manival torrent (Massif de la Chartreuse, French Prealps). A Light Detecting and Ranging (LiDAR) generated Digital Elevation Model (DEM) was used to identify five abandoned channels and related depositional forms (lobes, lateral levees) in the proximal alluvial fan of the torrent. A total of 156 Scots pine trees (Pinus sylvestris L.) with clear signs of debris flow events was analyzed and growth disturbances (GD) assessed, such as callus tissue, the onset of compression wood or abrupt growth suppression. In total, 375 GD were identified in the tree-ring samples, pointing to 13 debris-flow events for the period 1931–2008. While debris flows appear to be very common at Manival, they have only rarely propagated outside the main channel over the past 80 years. Furthermore, analysis of the spatial distribution of disturbed trees contributed to the identification of four patterns of debris-flow routing and led to the determination of three preferential breakout locations. Finally, the results of this study demonstrate that the temporal distribution of debris flows did not exhibit significant variations since the beginning of the 20th century.

Periosteal BMP2 activity drives bone graft healing

Bone graft incorporation depends on the orchestrated activation of numerous growth factors and cytokines in both the host and the graft. Prominent in this signaling cascade is BMP2. Although BMP2 is dispensable for bone formation, it is required for the initiation of bone repair; thus understanding the cellular mechanisms underlying bone regeneration driven by BMP2 is essential for improving bone graft therapies. In the present study, we assessed the role of Bmp2 in bone graft incorporation using mice in which Bmp2 has been removed from the limb prior to skeletal formation (Bmp2(cKO)). When autograft transplantations were performed in Bmp2cKO mice, callus formation and bone healing were absent. Transplantation of either a vital wild type (WT) bone graft into a Bmp2(cKO) host or a vital Bmp2(cKO) graft into a WT host also resulted in the inhibition of bone graft incorporation. Histological analyses of these transplants show that in the absence of BMP2, periosteal progenitors remain quiescent and healing is not initiated. When we analyzed the expression of Sox9, a marker of chondrogenesis, on the graft surface, we found it significantly reduced when BMP2 was absent in either the graft itself or the host, suggesting that local BMP2 levels drive periosteal cell condensation and subsequent callus cell differentiation. The lack of integrated healing in the absence of BMP2 was not due to the inability of periosteal cells to respond to BMP2. Healing was achieved when grafts were pre-soaked in rhBMP2 protein, indicating that periosteal progenitors remain responsive in the absence of BMP2. In contrast to the requirement for BMP2 in periosteal progenitor activation in vital bone grafts, we found that bone matrix-derived BMP2 does not significantly enhance bone graft incorporation. Taken together, our data show that BMP2 signaling is not essential for the maintenance of periosteal progenitors, but is required for the activation of these progenitors and their subsequent differentiation along the osteo-chondrogenic pathway. These results indicate that BMP2 will be among the signaling molecules whose presence will determine success or failure of new bone graft strategies.

Expression of antagonists of WNT and BMP signaling after non-rigid fixation of osteotomies

Delayed fracture healing and non-unions represent rare but severe complications in orthopedic surgery. Further knowledge on the mechanisms of the bone repair process and of the development of a pseudoarthrosis is essential to predict and prevent impaired healing of fractures. The present study aimed at elucidating differences in gene expression during the repair of rigidly and non-rigidly fixed osteotomies. For this purpose, the MouseFix™ and the FlexiPlate™ systems (AO Development Institute, Davos, CH), allowing the creation of well defined osteotomies in mouse femora, were employed. A time course following the healing process of the osteotomy was performed and bones and periimplant tissues were analyzed by high-resolution X-ray, MicroCT and by histology. For the assessment of gene expression, Low Density Arrays (LDA) were done. In animals with rigid fixation, X-ray and MicroCT revealed healing of the osteotomy within 3 weeks. Using the FlexiPlate™ system, the osteotomy was still visible by X-ray after 3 weeks and a stabilizing cartilaginous callus was formed. After 4.5 weeks, the callus was remodeled and the osteotomy was, on a histological level, healed. Gene expression studies revealed levels of transcripts encoding proteins associated with inflammatory processes not to be altered in tissues from bones with rigid and non-rigid fixation, respectively. Levels of transcripts encoding proteins of the extracellular matrix and essential for bone cell functions were not increased in the rigidly fixed group when compared to controls without osteotomy. In the FlexiPlate™ group, levels of transcripts encoding the same set of genes were significantly increased 3 weeks after surgery. Expression of transcripts encoding BMPs and BMP antagonists was increased after 3 weeks in repair tissues from bones fixed with FlexiPlate™, as were inhibitors of the WNT signaling pathways. Little changes only were detected in transcript levels of tissues from rigidly fixed bones. The data of the present study suggest that rigid fixation enables accelerated healing of an experimental osteotomy as compared to non-rigid fixation. The changes in the healing process after non-rigid fixation are accompanied by an increase in the levels of transcripts encoding inhibitors of osteogenic pathways and, probably as a consequence, by temporal changes in bone matrix synthesis.

Clinical evaluation of the CRIF 4.5/5.5 system for long-bone fracture repair in cattle

OBJECTIVE: To report clinical evaluation of the clamp rod internal fixator 4.5/5.5 (CRIF 4.5/5.5) in bovine long-bone fracture repair. STUDY DESIGN: Retrospective study. ANIMALS: Cattle (n=22) with long-bone fractures. METHODS: Records for cattle with long-bone fractures repaired between 1999 and 2004 with CRIF 4.5/5.5 were reviewed. Quality of fracture repair, fracture healing, and clinical outcome were investigated by means of clinical examination, medical records, radiographs, and telephone questionnaire. RESULTS: Successful long-term outcome was achieved in 18 cattle (82%); 4 were euthanatized 2-14 days postoperatively because of fracture breakdowns. Two cattle had movement of clamps on the rod. Moderate to severe callus formation was evident in 11 cattle 6 months postoperatively. CONCLUSIONS: Movement of clamps on the rod was recognized as implant failure unique to the CRIF. This occurred in cattle with poor fracture stability because of an extensive cortical defect. The CRIF system may not be ideal to treat metacarpal/metatarsal fractures because its voluminous size makes skin closure difficult, thereby increasing the risk of postoperative infections. CLINICAL RELEVANCE: CRIF cannot be recommended for repair of complicated long-bone fractures in cattle.

Treatment of tibial fractures with plates using minimally invasive percutaneous osteosynthesis in dogs and cats

OBJECTIVES: The aim of the here described case series was to develop and evaluate the minimally invasive percutaneous osteosynthesis for the plate fixation of tibial fractures in dogs and cats. METHODS: Six dogs and four cats with shaft fractures of the tibia were treated using minimally invasive percutaneous osteosynthesis. Follow-up radiographs four to six weeks after fracture fixation were evaluated for fracture healing. For the long-term follow-up (minimum 2.4 years), owners were contacted by phone to complete a questionnaire. RESULTS: All fractures healed without the need for a second procedure. Follow-up radiographs obtained after four to six weeks in seven cases showed advanced bony healing with callus formation and filling of the fracture gaps with calcified tissue in all seven. All the patients had a good to excellent long-term result with full limb function. The time needed for regaining full limb use was two to three months. CLINICAL SIGNIFICANCE: Minimally invasive percutaneous osteosynthesis seems to be a useful technique for the treatment of tibial shaft fractures in dogs and cats.

Long-term reaction to bone cement in osteoporotic bone: new bone formation in vertebral bodies after vertebroplasty

Elderly patients frequently suffer from osteoporotic vertebral fractures resulting in the need of vertebroplasty or kyphoplasty. Nevertheless, no data are available about the long-term consequences of cement injection into osteoporotic bone. Therefore, the aim of the present study was to evaluate the long-term tissue reaction on bone cement injected to osteoporotic bone during vertebroplasty. The thoracic spine of an 80-year-old female was explanted 3.5 years after vertebroplasty with polymethylmethacrylate. The treatment had been performed due to painful osteoporotic compression fractures. Individual vertebral bodies were cut in axial or sagittal sections after embedding. The sections were analysed using contact radiography and staining with toluidine blue. Furthermore, selected samples were evaluated with scanning electron microscopy and micro-compted tomography (in-plane resolution 6 microm). Large amounts of newly formed callus surrounding the injected polymethylmethacrylate were detected with all imaging techniques. The callus formation almost completely filled the spaces between the vertebral endplate, the cancellous bone, and the injected polymethylmethacrylate. In trabecular bone microfractures and osteoclast lacuna were bridged or filled with newly formed bone. Nevertheless, the majority of the callus formation was found in the immediate vicinity of the polymethylmethacrylate without any obvious relationship to trabecular fractures. The results indicate for the first time that, contrary to established knowledge, even in osteoporosis the formation of large amounts of new bone is possible.

Neuromuscular issues in cavovarus foot

Differential muscle weakness can cause a cavus foot deformity. Presenting complaints in the hindfoot may include ankle instability, secondary arthritis, or peroneal tendonitis. Presenting complaints in the forefoot may include stress fractures, callus formation over the lateral border of the foot, claw toes, first ray overload, and metatarsalgia. More general presenting complaints can include a drop-foot gait, decreased walking tolerance, and difficulty with shoe or orthotic fitting. To surgically correct the foot shape, soft tissue contractures need to be released, bone deformity corrected, and muscles balanced to optimize their strength and prevent recurrence of the deformity. This article reviews the diagnosis and management of the cavovarus foot secondary to longstanding muscle imbalance.

Puma indirectly activates Bax to cause apoptosis in the absence of Bid or Bim

Bcl-2 family members regulate apoptosis in response to cytokine withdrawal and a broad range of cytotoxic stimuli. Pro-apoptotic Bcl-2 family members Bax and Bak are essential for apoptosis triggered by interleukin-3 (IL-3) withdrawal in myeloid cells. The BH3-only protein Puma is critical for initiation of IL-3 withdrawal-induced apoptosis, because IL-3-deprived Puma(-/-) cells show increased capacity to form colonies when IL-3 is restored. To investigate the mechanisms of Puma-induced apoptosis and the interactions between Puma and other Bcl-2 family members, we expressed Puma under an inducible promoter in cells lacking one or more Bcl-2 family members. Puma rapidly induced apoptosis in cells lacking the BH3-only proteins, Bid and Bim. Puma expression resulted in activation of Bax, but Puma killing was not dependent on Bax or Bak alone as Puma readily induced apoptosis in cells lacking either of these proteins, but could not kill cells deficient for both. Puma co-immunoprecipitated with the anti-apoptotic Bcl-2 family members Bcl-x(L) and Mcl-1 but not with Bax or Bak. These data indicate that Puma functions, in the context of induced overexpression or IL-3 deprivation, primarily by binding and inactivating anti-apoptotic Bcl-2 family members.

The influence of different osteosynthesis configurations with locking compression plates (LCP) on stability and fracture healing after an oblique 45° angle osteotomy

Background Locking compression plates are used in various configurations with lack of detailed information on consequent bone healing. Study design In this in vivo study in sheep 5 different applications of locking compression plate (LCP) were tested using a 45° oblique osteotomy simulating simple fracture pattern. 60 Swiss Alpine sheep where assigned to 5 different groups with 12 sheep each (Group 1: interfragmentary lag screw and an LCP fixed with standard cortex screws as neutralisation plate; Group 2: interfragmentary lag screw and LCP with locking head screws; Group 3: compression plate technique (hybrid construct); Group 4: internal fixator without fracture gap; Group 5: internal fixator with 3 mm gap at the osteotomy site). One half of each group (6 sheep) was monitored for 6 weeks, and the other half (6 sheep) where followed for 12 weeks. Methods X-rays at 3, 6, 9 and 12 weeks were performed to monitor the healing process. After sacrifice operated tibiae were tested biomechanically for nondestructive torsion and compared to the tibia of the healthy opposite side. After testing specimens were processed for microradiography, histology, histomorphometry and assessment of calcium deposition by fluorescence microscopy. Results In all groups bone healing occurred without complications. Stiffness in biomechanical testing showed a tendency for higher values in G2 but results were not statistically significant. Values for G5 were significantly lower after 6 weeks, but after 12 weeks values had improved to comparable results. For all groups, except G3, stiffness values improved between 6 and 12 weeks. Histomorphometrical data demonstrate endosteal callus to be more marked in G2 at 6 weeks. Discussion and conclusion All five configurations resulted in undisturbed bone healing and are considered safe for clinical application.

Heritabilities of health traits in Swiss Warmblood horses

REASONS FOR PERFORMING THE STUDY: There is a lack of evidence regarding genetic parameters of health traits in Swiss Warmblood horses. OBJECTIVES: To estimate heritabilities of equine sarcoid disease, horn quality of the hooves, prognathism and increased filling of talocrural joints as a possible indicator for osteochondrosis in Swiss Warmblood horses examined at the field tests for 3-year-olds between 2005 and 2013. STUDY DESIGN: Retrospective analysis of breed society database. METHODS: Swiss Warmblood horses were examined clinically by 13 veterinarians at field tests in Switzerland between 2005 and 2013. The presence of sarcoids, horn quality of the hooves, incisor occlusion and increased joint filling were assessed and recorded. Records of 3715 horses were integrated in a pedigree comprising 217,282 horses. Variance components and heritabilities were estimated on the liability scale using MTGSAM. RESULTS: The prevalences of the examined traits were rather low, ranging from 2.4 to 13.0%. Heritabilities estimated were 0.21 ± 0.07 for the occurrence of sarcoids, 0.04 ± 0.02 for hooves with markedly brittle and friable horn quality, 0.03 ± 0.01 for hooves with marked growth ring formation, 0.06 ± 0.03 for prognathism and 0.08 ± 0.04 for increased filling of the talocrural joint (an indicator of possible osteochondrosis). The influence of the examiner on the variance of these observations was considerable. CONCLUSIONS: With the exception of equine sarcoid disease, estimates for the heritabilities for the traits examined here were low. A standardised examination protocol may reduce the variance due to the examiner. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Micropropagación de Cissus tiliacea, planta del sur del estado de México

Se micropropagó Cissus tiliacea, recurso fitogenético con potencial agronómico y farmacológico, en los medios de cultivo Murashige-Skoog (MS) y Lloyd y McCown (WPM). En ambos medios se generaron resultados similares para número de brotes, nudos, hojas y raíces adventicias, sólo existió diferencia significativa (p ≤ 0,05) en la formación de callo. Para la multiplicación in vitro se utilizó WPM adicionado con 0; 0,5; 1,0; 1,5 ó 2,0 mg L-1 de benciladenina (BA) y se emplearon tres tipos de segmentos nodales (basal, medio y apical). Las concentraciones de 0 y 0,5 mg L-1 de BA resultaron en un mayor tamaño y desarrollo del explante, además permitieron la formación de 1,2 a 1,6 raíces por explante. Las concentraciones de 1,5 y 2,0 mg L-1 de BA indujeron la formación de callo. No existió diferencia significativa en las variables evaluadas por efecto del tipo de segmento nodal establecido in vitro. En el enraizamiento, en el medio MS, se evaluaron tres tipos de auxinas: ácido naftalen-1-acético (ANA), ácido indol-3-butírico (AIB) y ácido indol- 3-acético (AIA) a 0,5 mg L-1; el mayor número de raíces secundarias y diámetro de la raíz principal fue inducido por ANA, sin embargo AIB indujo una mayor elongación de la raíz principal. Los resultados del presente trabajo sugieren que el cultivo in vitro de C. tiliacea es una alternativa para su conservación y multiplicación.

In vitro propagation of Glandularia peruviana (L.) Small, an ornamental native plant from South America

The flower market is characterized by being both eager for novelties and highly competitive. The exploration of native species with ornamental potential represents a remarkable area of research, since it entails the introduction and development of novel promising ornamental crops. The genus Glandularia, widely distributed in Argentina, holds an enormous ornamental potential, due to the variety of colors of its inflorescences (red, violet, white, rose and lily), and extended flowering period. There is little information on tissue culture of Glandularia, thus highlighting the relevance of this research. In this work, the conditions for in vitro multiplication of G. peruviana were optimized. It was concluded that WPM supplemented with TDZ, in concentrations ranging from 1.1 to 9.0 μM, was the most adequate treatment, rendering a multiplication rate of approximately 10 de novo shoots per explant. This paper presents a protocol for the in vitro propagation of this species and introduces interesting prospects in the application of biotechnological tools to breed Glandularia.

Isotopic ratios, concentration of noble gases and mineral and bulk composition of extraterrestrial dust particles in Dome Fuji ice core, East Antarctica

Two silicate-rich dust layers were found in the Dome Fuji ice core in East Antarctica, at Marine Isotope Stages 12 and 13. Morphologies, textures, and chemical compositions of constituent particles reveal that they are high-temperature melting products and are of extraterrestrial origin. Because similar layers were found ~2000 km east of Dome Fuji, at EPICA (European Project for Ice Coring in Antarctica)-Dome C, particles must have rained down over a wide area 434 and 481 ka. The strewn fields occurred over an area of at least 3 × 10**6 km**2. Chemical compositions of constituent phases and oxygen isotopic composition of olivines suggest that the upper dust layer was produced by a high-temperature interaction between silicate-rich melt and water vapor due to an impact explosion or an aerial burst of a chondritic meteoroid on the inland East Antarctic ice sheet. An estimated total mass of the impactor, on the basis of particle flux and distribution area, is at least 3 × 10**9 kg. A possible parent material of the lower dust layer is a fragment of friable primitive asteroid or comet. A hypervelocity impact of asteroidal/cometary material on the upper atmosphere and an explosion might have produced aggregates of sub-µm to µm-sized spherules. Total mass of the parent material of the lower layer must exceed 1 × 10**9 kg. The two extraterrestrial horizons, each a few millimeters in thickness, represent regional or global meteoritic events not identified previously in the Southern Hemisphere.

Chemistry of sulfide deposits from ODP Site 106-649

In Snake Pit massive sulfide fragments and friable, unconsolidated material recovered during ODP Leg 106, isocubanite and pyrite are generally the predominant phases, followed by marcasite, chalcopyrite, sphalerite, and pyrrhotite. Detailed analyses of paragenetic relations of minerals indicate that isocubanite first precipitated together with pyrrhotite. With decreasing temperature, chalcopyrite and sphalerite precipitated, and at the latest stage colloform sphalerite-pyrite (or colloform marcasite) formed. Isocubanite usually has exsolution lamellae of chalcopyrite and less commonly of pyrrhotite. The average bulk chemical composition of the friable, unconsolidated material indicates that it is rich in copper, reflecting the dominance of isocubanite in the specimens, and is characterized by high Co, low Pb, and Ag contents. Sulfur isotope ratios are very uniform, ranging in d34S from +1.2 to +2.8 per mil. The obtained values are apparently low, compared to those for the eastern Pacific sulfide samples, reflecting a smaller contribution of seawater sulfate in the Snake Pit sulfide deposit.