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71.
Apoptosis is characterized by DNA strand breaks with a 3'-OH terminus, which are analyzed by terminal deoxy(d)-UTP nick end labeling (TUNEL). Proteinase K digestion is thought to be an essential step in the TUNEL procedure. The effects of decalcifying reagents on general staining and the TUNEL assay for cartilage sections are largely unknown. The effects of these reagents on retention and integrity of DNA in chondrocytes have not been described until now. We evaluated the effects of various decalcifying solutions, including 10% EDTA, 10% citric acid, 5% trichloroacetic acid, 5% acetic acid and a commercial hydrochloric acid-based reagent, on general cartilage staining and the TUNEL assay for cartilage. The effects of proteinase K on nucleus preservation were also examined. Decalcification with 10% EDTA gave the best result for general cartilage staining. Chondrocyte DNA was retained and intact after using this reagent. Decalcification with 10% EDTA is also the safest method of decalcification if the TUNEL assay is applied to cartilage. Proteinase K digestion may have adverse effects on nucleus preservation in cartilage. Awareness of these effects is important whenever the TUNEL assay is applied. 相似文献
72.
MG Mullender NA Blom M De Kleuver JM Fock WMGC Hitters AMC Horemans CJ Kalkman JEH Pruijs RR Timmer PJ Titarsolej NC Van Haasteren Tol-de MJ Van Jager AJ Van Vught BJ Van Royen 《Scoliosis》2008,3(1):1-14
Background
Children with neuromuscular disorders with a progressive muscle weakness such as Duchenne Muscular Dystrophy and Spinal Muscular Atrophy frequently develop a progressive scoliosis. A severe scoliosis compromises respiratory function and makes sitting more difficult. Spinal surgery is considered the primary treatment option for correcting severe scoliosis in neuromuscular disorders. Surgery in this population requires a multidisciplinary approach, careful planning, dedicated surgical procedures, and specialized after care.Methods
The guideline is based on scientific evidence and expert opinions. A multidisciplinary working group representing experts from all relevant specialties performed the research. A literature search was conducted to collect scientific evidence in answer to specific questions posed by the working group. Literature was classified according to the level of evidence.Results
For most aspects of the treatment scientific evidence is scarce and only low level cohort studies were found. Nevertheless, a high degree of consensus was reached about the management of patients with scoliosis in neuromuscular disorders. This was translated into a set of recommendations, which are now officially accepted as a general guideline in the Netherlands.Conclusion
In order to optimize the treatment for scoliosis in neuromuscular disorders a Dutch guideline has been composed. This evidence-based, multidisciplinary guideline addresses conservative treatment, the preoperative, perioperative, and postoperative care of scoliosis in neuromuscular disorders. 相似文献73.
The main advances in immunology have been forged or underpinned by animal experiments. However, animal research now focuses excessively on one laboratory species, and there is too much redundant repetition and too few transfers from basic discovery to successful clinical application. These features can be improved markedly by placing more emphasis on biological relevance when evaluating animal models and by taking greater advantage of the unique experimental opportunities that are offered by large animals. 相似文献
74.
PJ?Baker H?Johnston M?Abel HM?Charlton PJ?O'ShaughnessyEmail author 《Reproductive biology and endocrinology : RB&E》2003,1(1):4
During mammalian testis development distinct generations of fetal and adult Leydig cells arise. Luteinising hormone (LH) is
required for normal adult Leydig cell function and for the establishment of normal adult Leydig cell number but its role in
the process of adult Leydig cell differentiation has remained uncertain. In this study we have examined adult Leydig cell
differentiation in gonadotrophin-releasing hormone (GnRH)-null mice which are deficient in circulating gonadotrophins. Adult
Leydig cell differentiation was assessed by measuring expression of mRNA species encoding four specific markers of adult Leydig
cell differentiation in the mouse. Each of these markers (3β-hydroxysteroid dehydrogenase type VI (3βHSD VI), 17β-hydroxysteroid
dehydrogenase type III (17βHSD III), prostaglandin D (PGD)-synthetase and oestrogen sulphotransferase (EST)) is expressed
only in the adult Leydig cell lineage in the normal adult animal. Real-time PCR studies showed that all four markers are expressed
in adult GnRH-null mice. Localisation of 3βHSD VI and PGD-synthetase expression by in situ hybridisation confirmed that these genes are expressed in the interstitial tissue of the GnRH-null mouse. Treatment of animals
with human chorionic gonadotrophin increased expression of 3βHSD VI and 17βHSD III within 12 hours further indicating that
differentiated, but unstimulated cells already exist in the GnRH-null mouse. Thus, while previous studies have shown that
LH is required for adult Leydig cell proliferation and activity, results from the present study show that adult Leydig cell
differentiation will take place in animals deficient in LH. 相似文献
75.
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77.
K. Evangelou J. Bramis I. Peros P. Zacharatos D. Dasiou-Plakida N. Kalogeropoulos PJ Asimacopoulos C. Kittas E. Marinos VG Gorgoulis 《Biotechnic & histochemistry》2004,79(1):5-10
It is well established that p16INK4A protein acts as a cell cycle inhibitor in the nucleus. Therefore, cytoplasmic localization of p16 INK4A usually is disregarded by investigators as nonspecific. Three recent studies reported findings that differ from the current view concerning p16INK4A immunohistochemical localization. All three demonstrated that breast and colon cancers expressing cytoplasmic p16INK4 represent distinct biological subsets. We previously detected in a percentage of non-small cell lung carcinomas simultaneous nuclear and cytoplasmic p16INK4A staining. In view of the reports concerning breast and colon carcinomas, we conducted an ultrastructural re-evaluation of our cases to clarify the specificity of p16INK4A cytoplasmic expression. We observed p16 INK4A immunolocalization in both the nucleus and the cytoplasm of a proportion of tumor cells. Diffuse dense nuclear staining was detected in the nucleoplasm, whereas weaker granular immunoreactivity was observed in the cytoplasm near the rough endoplasmic reticulum. Negative tumor cells also were visible. In the tumor-associated stromal, cells p16INK4A immunoreactivity was detected only in the nuclei. We have demonstrated that p16INK4A cytoplasmic staining is specific and suggest that it represents a mechanism of p16INK4A inactivation similar to that observed in other tumor suppressor genes. 相似文献
78.
79.
Xavier Bailly Elisa Giuntini M Connor Sexton Ryan PJ Lower Peter W Harrison Nitin Kumar J Peter W Young 《The ISME journal》2011,5(11):1722-1734
We investigated the genomic diversity of a local population of the symbiotic bacterium Sinorhizobium medicae, isolated from the roots of wild Medicago lupulina plants, in order to assess genomic diversity, to identify genomic regions influenced by duplication, deletion or strong selection, and to explore the composition of the pan-genome. Partial genome sequences of 12 isolates were obtained by Roche 454 shotgun sequencing (average 5.3 Mb per isolate) and compared with the published sequence of S. medicae WSM 419. Homologous recombination appears to have less impact on the polymorphism patterns of the chromosome than on the chromid pSMED01 and megaplasmid pSMED02. Moreover, pSMED02 is a hot spot of insertions and deletions. The whole chromosome is characterized by low sequence polymorphism, consistent with the high density of housekeeping genes. Similarly, the level of polymorphism of symbiosis genes (low) and of genes involved in polysaccharide synthesis (high) may reflect different selection. Finally, some isolates carry genes that may confer adaptations that S. medicae WSM 419 lacks, including homologues of genes encoding rhizobitoxine synthesis, iron uptake, response to autoinducer-2, and synthesis of distinct polysaccharides. The presence or absence of these genes was confirmed by PCR in each of these 12 isolates and a further 27 isolates from the same population. All isolates had rhizobitoxine genes, while the other genes were co-distributed, suggesting that they may be on the same mobile element. These results are discussed in relation to the ecology of Medicago symbionts and in the perspective of population genomics studies. 相似文献
80.
MOTIVATION: The first version of the graphical multiple sequence alignment environment QAlign was published in 2003. Heavy response from the molecular-biological user community clearly demonstrated the need for such a platform. RESULTS: Panta rhei extends QAlign by several features. Major redesigns on the user interface, for instance, allow users to flexibily compose views for multiple projects. The new sequence viewer handles datasets with arbitrarily many and arbitrarily large sequences that may still be edited by guided block moving. More distance-based algorithms are available to interactively reconstruct phylogenetic trees which can now also be zoomed and navigated graphicaly. AVAILABILITY: Executables and the JAVA source code are available under the Apache license at http://gi.cebitec.uni-bielefeld.de/qalign CONTACT: qalign@cebitec.uni-bielefeld.de. 相似文献