The Rare Knowledge Mining Methodological Framework for the Development of Practice Guidelines and Knowledge Translation Tools for Rare Diseases.

Rare diseases bring on a heavy health, social and economic burden that impacts patients' lives and puts pressure on the healthcare system. Furthermore, they are often associated with limited published studies to inform multidisciplinary clinical practice thus limiting evidence-based practice. Moreover, the development of knowledge translation products including clinical care guidelines are often very challenging based on the current available methodological frameworks relying mostly on critical appraisal of the published research evidence where randomized clinical trial design is considered as the gold standard. To overcome this barrier, we proposed the Rare Knowledge Mining Methodological Framework (RKMMF). The RKMMF is one possible answer to improve the development of knowledge translation products for rare diseases. This framework includes other sources of evidence including registry information and qualitative studies and the involvement of expert patients. This article documents the RKMMF structure and its application is exemplified through knowledge translation products developed for a neuromuscular population.

Differential sensitivity to apoptosis between the human small and large intestinal mucosae: linkage with segment-specific regulation of BCL-2 homologs and involvement of signaling pathways.

The small and large intestines differ in their expression profiles of Bcl-2 homologs. Intestinal segment-specific Bcl-2 homolog expression profiles are acquired as early as by mid-gestation (18-20 weeks) in man. In the present study, we examined the question whether such distinctions underlie segment-specific control mechanisms of intestinal cell survival. Using mid-gestation human jejunum and colon organotypic cultures, we analyzed the impact of growth factors (namely insulin; 10 microg/ml) and pharmacological compounds that inhibit signal transduction molecules/pathways (namely tyrosine kinases, Fak, P13-K/Akt, and MEK/Erk) on cell survival and Bcl-2 homolog expression (anti-apoptotic: Bcl-2, Bcl-X(L), Mcl-1; pro-apoptotic: Bax, Bak, Bad). The relative activation levels of p125Fak, p42Erk-2, and p57Akt were analyzed as well. Herein, we report that (1) the inhibition of signal transduction molecules/pathways revealed striking differences in their impact on cell survival in the jejunum and colon (e.g., the inhibition of p125Fak induced apoptosis with a significantly greater extent in the jejunum [approximately 43%] than in the colon [approximately 24%]); (2) sharp distinctions between the two segments were noted in the modulatory effects of the various treatments on Bcl-2 homolog steady-state levels (e.g., inhibition of tyrosine kinase activities in the jejunum down-regulated all anti-apoptotics analyzed while increasing Bax, whereas the same treatment in the colon down-regulated Bcl-X(L) only and increased all pro-apoptotics); and (3) in addition to their differential impact on cell survival and Bcl-2 homolog expression, the MEK/Erk and P13-K/Akt pathways were found to be distinctively regulated in the jejunum and colon mucosae (e.g., insulin in the jejunum increased p42Erk-2 activation without affecting that of p57Akt, whereas the same treatment in the colon decreased p42Erk-2 activation while increasing that of p57Akt). Altogether, these data show that intestinal cell survival is characterized by segment-specific susceptibilities to apoptosis, which are in turn linked with segmental distinctions in the involvement of signaling pathways and the regulation of Bcl-2 homolog steady-state levels. Therefore, these indicate that cell survival is subject to segment-specific control mechanisms along the proximal-distal axis of the intestine.

Early acquisition of bowel segment-specific Bcl-2 homolog expression profiles during development of the human ileum and colon.

The adult small and large intestines display distinct expression profiles of Bcl-2 homologs, known regulators of apoptosis. This is thought to indicate that control mechanisms of intestinal apoptosis are gut segment-specific. Little is known on the expression of Bcl-2 homologs during gut development. In man, intestinal features and functions are acquired largely by mid-gestation (18-20 wks); the question whether segment-specific controls of intestinal apoptosis are also acquired early during development remains open. In the present study, we approached this by investigating the expression of six Bcl-2 homologs (Bcl-2, Bcl-XL, Mcl-1, Bax, Bak, Bad), and one nonhomologous associated molecule (Bag-1), during development of the human ileum and colon (12-20 wks of gestation). Beginning at 18 wks, we found that the epithelial localization of Bcl-2 homologs displayed differential patterns (or gradients) in both the ileum and colon; however, the patterns of some of the homologs differed between the two segments. For instance, Bag-1 and Bcl-2 exhibited crypt-villus decreasing gradients of expression in the ileum but not in the colon, whereas Mcl-1 displayed differing compartimentalizations between the two segments. Further analyses indicated that the steady-state expression levels of Bcl-2 homologs underwent modulations between 12 and 20 wks; however, the observed developmental profiles contrasted significantly between the two segments. For example, Bcl-2, Bag-1 and Bak levels increased in the colon, but the levels of these same homologs decreased in the ileum. Furthermore, by 18-20 wks, we found that the expression levels of each Bcl-2 homolog analyzed differed greatly between the ileum and colon. Altogether, these data indicate that the expression of Bcl-2 homologs is modulated differentially during human gut development in order to establish, by mid-gestation, distinct expression profiles for the small and large intestines. This in turn suggests that gut segment-specific control mechanisms of human intestinal apoptosis are acquired early during fetal life.

Genetic diversity in populations of Cronartium ribicola in plantations and natural stands of Pinus strobus.

Genetic diversity was studied in 22 populations of the white pine blister rust fungus Cronartium ribicola from natural stands and plantations of eastern white pine, Pinus strobus. Pseudo-allelic frequencies were estimated at each of 7 putative RAPD loci by scoring for presence or absence of amplified fragments in dikaryotic aecidiospores. Analysis of genetic distance between all pairs of populations did not reveal any trend with regard to geographic origin or type of white pine stand. In addition, when hierarchical population structure was analysed, total genetic diversity (H s =0.214) was mostly attributable to diversity within populations (H s =0.199; AMOVA φ st =0.121, P<0.01). Genetic diversity of populations relative to region of origin (east, centre, and west) or type of stand (natural stands vs plantations) was not significantly different from zero (P>0.10) Nevertheless, a significant proportion of genetic differentiation was found between populations within region or stand type (F st =0.114; φ sc =0.132, P<0.001). This result indicates that some population structure exists but that it appears to be independent of region of origin or type of stand. At least for 2 populations from white pine plantations, it appears possible that a recent introduction of a limited number of propagules was responsible for low levels of genetic diversity. We interpret these results as meaning that either long-distance dispersal is taking place between populations more than 1000 km apart or that these populations share a common recent ancestor. In addition, we suggest that C. ribicola may still be expanding its distribution by colonizing new plantations.

A novel intergeneric hybrid in the Triticeae: Triticum aestivum x Psathyrost achys juncea.

Two hybrid embryos of intergeneric origin between Triticum aestivum cv Fukuho (2n=6x=42, AABBDD) and Psathyrostachys juncea (2n=2x=14, NN) were successfully rescued. One hybrid plant had the expected chromosome number of 28 (ABDN), whereas the second plant had 35 chromosomes. The average meiotic chromosome pairing in the 35-chromosome hybrid was 21.87 univalents + 6.38 bivalents + 0.11 trivalents + 0.009 quadrivalents, which indicates that two copies of the N genome were present. Chromosome pairing in the 28-chromosome hybrid was low (1.35 bivalents), and pointed out the lack of homology between the wheat genomes and the P. juncea genome. These new hybrids showed some necrosis and chlorosis, which caused severe floral abortion in the plant that had 35 chromosomes. These problems became gradually less severe after 18 months.

Somatic embryogenesis and plant regeneration in Triticum aestivum x Leymus angustus F1 hybrids and the parental lines.

Somatic embryos and plants were produced from cultured inflorescence and leaf segments of Triticum aestivum X Leymus anaustus F1 hybrids and the parental lines. Inflorescences showed a better capacity for somatic embryogenesis and plant regeneration than leaves. Leymus anaustus produced the highest number of embryogenic calli, while the hybrids were intermediate between this species and Triticum aestivum. Presence of 2,4-D was shown to be essential for induction and maintenance of somatic embryogenesis. Addition of five amino acids (glutamine, proline, asparagine, aspartic acid and glutamic acid) did not have any marked effect when they were used in the callus induction medium. The regenerated plants had the same morphology as the original plants. No cytological modification was observed in the examined plants.

Production and cytogenetics of hybrids of Triticum aestivum x Leymus innovatus.

Hybrid plants were obtained between Triticum aestivum (2n=6x=42, AABBDD) and Leymus innovatus (2n=4x=28, JJNN) at a frequency varying from 0.4% to 1.2% of the pollinated florets. Improvement of the embryo culture medium resulted in a higher frequency of embryo rescue. Eight of ten hybrids had the expected chromosome number of 35 (ABDJN). Meiotic analysis indicated that there was no homology between the genomes of the two species. Two hybrids had only 28 chromosomes. Comparison of chromosome pairing between the two types of hybrids suggested that Leymus innovatus carries genes that affect chromosome pairing and behavior. The relatively high occurrence of spontaneous doubling in the meiocytes of these hybrids may indicate that backcrossing of the hybrids to wheat should be possible, although frequent chromosome irregularities observed in the meiocytes of the hybrids may decrease the probability of success of this step, which is essential to the process of gene transfer from L. innovatus to wheat.