A musculoskeletal shoulder simulation of moment arms and joint reaction forces after medialization of the supraspinatus footprint in rotator cuff repair

A non-anatomical reinsertion of the supraspinatus medially to the original footprint to avoid over-tensioning of the tendon in large and retracted tears is one surgical option in rotator cuff (RC) repair. The purpose of the study was to determine the biomechanical effects on the glenohumeral joint with regard to this surgical technique. A modified musculoskeletal computational shoulder model was used to evaluate the change in moment arms and muscle forces of the RC and the co-contracting muscles and the alteration of the joint reaction forces (compressive and shear forces) after reinsertion of the supraspinatus 5?mm, 10?mm, 15?mm and 20?mm medially to the original footprint. A medialization of the supraspinatus reduces its moment arm in glenohumeral abduction. In case of a medialization of the attachment of 15?mm and 20?mm, the supraspinatus restricts glenohumeral abduction at 54° and 68°. In glenohumeral forward flexion and in lower degrees of internal rotation the moment arm of the supraspinatus increases for a medialized tendon attachment and decreases in external rotation in relation to the anatomical condition. A medialization of the supraspinatus insertion point yields in an increase in muscle force for abduction, internal and external rotation. In the present model a medially non-anatomic reinsertion reduces significantly the compressive glenohumeral joint reaction and the glenohumeral stability. Moreover, the results show that a medialization of the supraspinatus leads to a reduction of the supraspinatus moment arm especially in abduction. This leads to an increase of a compensatory supraspinatus load for stabilization the humerus in space, which may potentially cause a postoperative overload of the tendon-bone-complex.     

Raman-FISH: combining stable-isotope Raman spectroscopy and fluorescence in situ hybridization for the single cell analysis of identity and function

We have coupled fluorescence in situ hybridization (FISH) with Raman microscopy for simultaneous cultivation-independent identification and determination of (13)C incorporation into microbial cells. Highly resolved Raman confocal spectra were generated for individual cells which were grown in minimal medium where the ratio of (13)C to (12)C content of the sole carbon source was incrementally varied. Cells which were (13)C-labelled through anabolic incorporation of the isotope exhibited key red-shifted spectral peaks, the calculated 'red shift ratio' (RSR) being highly correlated with the (13)C-content of the cells. Subsequently, Raman instrumentation and FISH protocols were optimized to allow combined epifluorescence and Raman imaging of Fluos, Cy3 and Cy5-labelled microbial populations at the single cell level. Cellular (13)C-content determinations exhibited good congruence between fresh cells and FISH hybridized cells indicating that spectral peaks, including phenylalanine resonance, which were used to determine (13)C-labelling, were preserved during fixation and hybridization. In order to demonstrate the suitability of this technology for structure-function analyses in complex microbial communities, Raman-FISH was deployed to show the importance of Pseudomonas populations during naphthalene degradation in groundwater microcosms. Raman-FISH extends and complements current technologies such as FISH-microautoradiography and stable isotope probing in that it can be applied at the resolution of single cells in complex communities, is quantitative if suitable calibrations are performed, can be used with stable isotopes and has analysis times of typically 1 min per cell.     

Synergistic effects of seasonal rainfall, parasites and demography on fluctuations in springbok body condition

1. Seasonality of rainfall can exert a strong influence on animal condition and on host-parasite interactions. The body condition of ruminants fluctuates seasonally in response to changes in energy requirements, foraging patterns and resource availability, and seasonal variation in parasite infections may further alter ruminant body condition. 2. This study disentangles the effects of rainfall and gastrointestinal parasite infections on springbok (Antidorcas marsupialis) body condition and determines how these factors vary among demographic groups. 3. Using data from four years and three study areas, we investigated (i) the influence of rainfall variation, demographic factors and parasite interactions on parasite prevalence or infection intensity, (ii) whether parasitism or rainfall is a more important predictor of springbok body condition and (iii) how parasitism and condition vary among study areas along a rainfall gradient. 4. We found that increased parasite intensity is associated with reduced body condition only for adult females. For all other demographic groups, body condition was significantly related to prior rainfall and not to parasitism. Rainfall lagged by two months had a positive effect on body condition. 5. Adult females showed evidence of a 'periparturient rise' in parasite intensity and had higher parasite intensity and lower body condition than adult males after parturition and during early lactation. After juveniles were weaned, adult females had lower parasite intensity than adult males. Sex differences in parasitism and condition may be due to differences between adult females and males in the seasonal timing of reproductive effort and its effects on host immunity, as well as documented sex differences in vulnerability to predation. 6. Our results highlight that parasites and the environment can synergistically affect host populations, but that these interactions might be masked by their interwoven relationships, their differential impacts on demographic groups, and the different time-scales at which they operate.     

Impact of polyunsaturated fatty acids on hepato-pancreatic prostaglandin and leukotriene concentration in ductal pancreatic cancer—Is there a correlation to tumour growth and liver metastasis?

Type and composition of polyunsaturated fatty acids (PUFAs) are suspected to play an important role in carcinogenesis. Thus we investigated the effects of n-3, n-6 and n-9 PUFAs on tumour growth, liver metastasis and concentration of prostaglandins (PG) and leukotrienes (LT) in experimental ductal pancreatic adenocarcinoma. Ninety male hamsters were randomised into six groups (Gr.) (n=15). While Gr. 1-3 were healthy control groups, Gr. 4-6 weekly received subcutaneous injections of 10mg N-nitrosobis-2-oxypropylamine (BOP)/kg body weight for 12 weeks in order to induce ductal pancreatic adenocarcinoma. Between week 1 and 16 all animals were fed with a standard diet with a raw fat content of 2.9%. In week 17 Gr. 1-6 were allocated to three types of diets: Gr. 1: standard high fat (=SHF diet, rich in n-6 PUFAs)/Gr. 2: FISH-OIL (rich in n-3 PUFAs)/Gr. 3: SMOF (=mixture of n-3, n-6 and n-9 PUFAs)/Gr. 4: BOP+SHF/Gr. 5: BOP+SMOF/Gr. 6: BOP+FISH-OIL. After 32 weeks all animals were sacrificed and pancreas as well as liver were analysed histologically. Furthermore pancreatic and hepatic concentrations of prostaglandins (PGF1alpha, PGE(2)) and LT were measured. FISH-OIL decreased number of macroscopically visible pancreatic tumours (Gr. 4-6: 54.5% vs. 45.5% vs. 9.1%, P<0.05) as well as incidence of liver metastasis (Gr. 4-6: 90.9% vs. 72.7% vs. 36.4%, P<0.05). Furthermore concentration of PGF(1)(alpha), PGE(2) and LT were significantly increased in pancreatic carcinoma compared to tumour-free tissue. Moreover levels of PGF(1)(alpha) and PGE(2) were higher in liver metastasis than in extrametastatic hepatic tissue. However, in Gr. 6 (FISH-OIL) intrametastatic concentration of LT was significantly lower than in non-metastatic hepatic tissue as well as in Gr. 4 and Gr. 5. FISH-OIL decreased number of visible pancreatic tumours and incidence of histological proven liver metastasis. This effect might be caused by a decrease of intrametastatic concentration of LT compared to extrametastatic hepatic tissue.     

Matrix metalloproteinase inhibitor RO 28-2653 decreases liver metastasis by reduction of MMP-2 and MMP-9 concentration in BOP-induced ductal pancreatic cancer in Syrian Hamsters: inhibition of matrix metalloproteinases in pancreatic cancer

BACKGROUND: Matrix metalloproteinases (MMP) are proteolytic enzymes which degrade the extracellular matrix and therefore play an important role in metastasis. However, the impact of MMP inhibitors (MMPI) on pancreatic cancer is still unclear. Thus we evaluated the influence of selective MMPI Ro 28-2653 on the incidence of liver metastases and the concentration of MMP-2 and MMP-9 in ductal pancreatic adenocarcinoma in Syrian hamster. MATERIAL AND METHODS: One hundred and thirty male Syrian hamsters were randomised into 8 groups (Gr.1-3: n=15, Gr.4-8: n=17). Pancreatic cancer was induced by weekly subcutaneous injection of 10mg N-nitrosobis-2-oxopropylamin (BOP)/kg body weight (Gr.4-8) while healthy control Gr. 1-3 received 0.5 ml sodium chloride 0.9%. Gr.1 and 4 had free access to a standard diet, Gr. 2, 3 and 5-8 received a diet rich in polyunsaturated fatty acids, which increases liver metastasis in this model. In week 17 oral therapy started: Gr.3 and 6: 60 mg Eudragit/kg body weight/d (vehicle of MMPI), Gr.7 and 8: 40 mg, respectively, 120 mg RO 28-2653/kg body weight/d; Gr.1, 2, 4, 5: no therapy. After 30 weeks all hamsters were sacrificed and histopathologically examined. Additionally concentrations of MMP-2 and MMP-9 were measured in non-metastatic liver and liver metastases. RESULTS: Concentrations of MMP-2 and MMP-9 in liver metastases were decreased by high- and low-dose therapy with MMPI. Furthermore, the incidence of liver metastases was significantly reduced by low-dose therapy with Ro 28-2653. CONCLUSION: Low-dose therapy with Ro 28-2653 decreased liver metastasis due to an inhibition of MMP-2 and MMP-9 concentration in ductal pancreatic cancer.     

Development and assessment of Diversity Arrays Technology for high-throughput DNA analyses in <Emphasis Type="Italic">Musa</Emphasis>

Diversity Arrays Technology (DArT) is a DNA hybridisation-based molecular marker technique that can detect simultaneously variation at numerous genomic loci without sequence information. This efficiency makes it a potential tool for a quick and powerful assessment of the structure of germplasm collections. This article demonstrates the usefulness of DArT markers for genetic diversity analyses of Musa spp. genotypes. We developed four complexity reduction methods to generate DArT genomic representations and we tested their performance using 48 reference Musa genotypes. For these four complexity reduction methods, DArT markers displayed high polymorphism information content. We selected the two methods which generated the most polymorphic genomic representations (PstI/BstNI 16.8%, PstI/TaqI 16.1%) to analyze a panel of 168 Musa genotypes from two of the most important field collections of Musa in the world: Cirad (Neufchateau, Guadeloupe), and IITA (Ibadan, Nigeria). Since most edible cultivars are derived from two wild species, Musa acuminata (A genome) and Musa balbisiana (B genome), the study is restricted mostly to accessions of these two species and those derived from them. The genomic origin of the markers can help resolving the pedigree of valuable genotypes of unknown origin. A total of 836 markers were identified and used for genotyping. Ten percent of them were specific to the A genome and enabled targeting this genome portion in relatedness analysis among diverse ploidy constitutions. DArT markers revealed genetic relationships among Musa genotype consistent with those provided by the other markers technologies, but at a significantly higher resolution and speed and reduced cost.     

Meiosis Drives Extraordinary Genome Plasticity in the Haploid Fungal Plant Pathogen Mycosphaerella graminicola

Meiosis in the haploid plant-pathogenic fungus Mycosphaerella graminicola results in eight ascospores due to a mitotic division following the two meiotic divisions. The transient diploid phase allows for recombination among homologous chromosomes. However, some chromosomes of M. graminicola lack homologs and do not pair during meiosis. Because these chromosomes are not present universally in the genome of the organism they can be considered to be dispensable. To analyze the meiotic transmission of unequal chromosome numbers, two segregating populations were generated by crossing genetically unrelated parent isolates originating from Algeria and The Netherlands that had pathogenicity towards durum or bread wheat, respectively. Detailed genetic analyses of these progenies using high-density mapping (1793 DArT, 258 AFLP and 25 SSR markers) and graphical genotyping revealed that M. graminicola has up to eight dispensable chromosomes, the highest number reported in filamentous fungi. These chromosomes vary from 0.39 to 0.77 Mb in size, and represent up to 38% of the chromosomal complement. Chromosome numbers among progeny isolates varied widely, with some progeny missing up to three chromosomes, while other strains were disomic for one or more chromosomes. Between 15–20% of the progeny isolates lacked one or more chromosomes that were present in both parents. The two high-density maps showed no recombination of dispensable chromosomes and hence, their meiotic processing may require distributive disjunction, a phenomenon that is rarely observed in fungi. The maps also enabled the identification of individual twin isolates from a single ascus that shared the same missing or doubled chromosomes indicating that the chromosomal polymorphisms were mitotically stable and originated from nondisjunction during the second division and, less frequently, during the first division of fungal meiosis. High genome plasticity could be among the strategies enabling this versatile pathogen to quickly overcome adverse biotic and abiotic conditions in wheat fields.     

Isolated chromosomes as a new and efficient source of DArT markers for the saturation of genetic maps

We describe how the diversity arrays technology (DArT) can be coupled with chromosome sorting to increase the density of genetic maps in specific genome regions. Chromosome 3B and the short arm of chromosome 1B (1BS) of wheat were isolated by flow cytometric sorting and used to develop chromosome- and chromosome arm-enriched genotyping arrays containing 2,688 3B clones and 384 1BS clones. Linkage analysis showed that 553 of the 711 polymorphic 3B-derived markers (78%) mapped to chromosome 3B, and 59 of the 68 polymorphic 1BS-derived markers (87%) mapped to chromosome 1BS, confirming the efficiency of the chromosome-sorting approach. To demonstrate the potential for saturation of genetic maps, we constructed a consensus map of chromosome 3B using 19 mapping populations, including some that were genotyped with the 3B-enriched array. The 3B-derived DArT markers doubled the number of genetic loci covered. The resulting consensus map, probably the densest genetic map of 3B available to this date, contains 939 markers (779 DArTs and 160 other markers) that segregate on 304 genetically distinct loci. Importantly, only 2,688 3B-derived clones (probes) had to be screened to obtain almost twice as many polymorphic 3B markers (510) as identified by screening approximately 70,000 whole genome-derived clones (269). Since an enriched DArT array can be developed from less than 5 ng of chromosomal DNA, a quantity which can be obtained within 1 h of sorting, this approach can be readily applied to any crop for which chromosome sorting is available.     

Low genetic variability and strong differentiation among isolated populations of the rare steppe grass Stipa capillata L. in Central Europe

Stipa capillata L. (Poaceae) is a rare grassland species in Central Europe that is thought to have once been widespread in post‐glacial times. Such relict species are expected to show low genetic diversity within populations and high genetic differentiation between populations due to bottlenecks, long‐term isolation and ongoing habitat fragmentation. These patterns should be particularly pronounced in selfing species. We analysed patterns of random amplified polymorphic DNA (RAPD) variation in the facultatively cleistogamous S. capillata to examine whether genetic diversity is associated with population size, and to draw initial conclusions on the migration history of this species in Central Europe. We analysed 31 S. capillata populations distributed in northeastern, central and western Germany, Switzerland and Slovakia. Estimates of genetic diversity at the population level were low and not related to population size. Among all populations, extraordinarily high levels of genetic differentiation (amova : φ_ST = 0.86; Bayesian analysis: θ^B = 0.758) and isolation‐by‐distance were detected. Hierarchical amova indicated that most of the variability was partitioned among geographic regions (59%), or among populations between regions when the genetically distinct Slovakian populations were excluded. These findings are supported by results of a multivariate ordination analysis. We also found two different groups in an UPGMA cluster analysis: one that contained the populations from Slovakia, and the other that combined the populations from Germany and Switzerland. Our findings imply that S. capillata is indeed a relict species that experienced strong bottlenecks in Central Europe, enhanced by isolation and selfing. Most likely, populations in Slovakia were not the main genetic source for the post‐glacial colonization of Central Europe.