Intramyocardial Transplantation and Tracking of Human Mesenchymal Stem Cells in a Novel Intra-Uterine Pre-Immune Fetal Sheep Myocardial Infarction Model: A Proof of Concept Study

Although stem-cell therapies have been suggested for cardiac-regeneration after myocardial-infarction (MI), key-questions regarding the in-vivo cell-fate remain unknown. While most available animal-models require immunosuppressive-therapy when applying human cells, the fetal-sheep being pre-immune until day 75 of gestation has been proposed for the in-vivo tracking of human cells after intra-peritoneal transplantation. We introduce a novel intra-uterine myocardial-infarction model to track human mesenchymal stem cells after direct intra-myocardial transplantation into the pre-immune fetal-sheep. Thirteen fetal-sheep (gestation age: 70–75 days) were included. Ten animals either received an intra-uterine induction of MI only (n = 4) or MI+intra-myocardial injection (IMI;n = 6) using micron-sized, iron-oxide (MPIO) labeled human mesenchymal stem cells either derived from the adipose-tissue (ATMSCs;n = 3) or the bone-marrow (BMMSCs;n = 3). Three animals received an intra-peritoneal injection (IPI;n = 3; ATMSCs;n = 2/BMMSCs;n = 1). All procedures were performed successfully and follow-up was 7–9 days. To assess human cell-fate, multimodal cell-tracking was performed via MRI and/or Micro-CT, Flow-Cytometry, PCR and immunohistochemistry. After IMI, MRI displayed an estimated amount of 1×10⁵–5×10⁵ human cells within ventricular-wall corresponding to the injection-sites which was further confirmed on Micro-CT. PCR and IHC verified intra-myocardial presence via detection of human-specific β-2-microglobulin, MHC-1, ALU-Sequence and anti-FITC targeting the fluorochrome-labeled part of the MPIOs. The cells appeared viable, integrated and were found in clusters or in the interstitial-spaces. Flow-Cytometry confirmed intra-myocardial presence, and showed further distribution within the spleen, lungs, kidneys and brain. Following IPI, MRI indicated the cells within the intra-peritoneal-cavity involving the liver and kidneys. Flow-Cytometry detected the cells within spleen, lungs, kidneys, thymus, bone-marrow and intra-peritoneal lavage, but not within the heart. For the first time we demonstrate the feasibility of intra-uterine, intra-myocardial stem-cell transplantation into the pre-immune fetal-sheep after MI. Utilizing cell-tracking strategies comprising advanced imaging-technologies and in-vitro tracking-tools, this novel model may serve as a unique platform to assess human cell-fate after intra-myocardial transplantation without the necessity of immunosuppressive-therapy.     

Crystal Structure of an Anti-Ang2 CrossFab Demonstrates Complete Structural and Functional Integrity of the Variable Domain

Bispecific antibodies are considered as a promising class of future biotherapeutic molecules. They comprise binding specificities for two different antigens, which may provide additive or synergistic modes of action. There is a wide variety of design alternatives for such bispecific antibodies, including the “CrossMab” format. CrossMabs contain a domain crossover in one of the antigen-binding (Fab) parts, together with the “knobs-and-holes” approach, to enforce the correct assembly of four different polypeptide chains into an IgG-like bispecific antibody. We determined the crystal structure of a hAng-2-binding Fab in its crossed and uncrossed form and show that C_H1-C_L-domain crossover does not induce significant perturbations of the structure and has no detectable influence on target binding.     

How life cycle–based science and practice support the transition towards a sustainable economy

The International Journal of Life Cycle Assessment -     

Movement and dispersal patterns in the bush cricket Pholidoptera griseoaptera: the role of developmental stage and sex

Abstract.  1. To persist in a highly fragmented environment, such as the intensively used agricultural landscapes of Europe, species may need to successfully disperse between sub-populations. This paper describes a capture – mark – resight experiment to investigate movement and dispersal patterns of the flightless bush cricket species Pholidoptera griseoaptera .
2. Movement parameters showed little difference between juveniles and imagos as well as between males and females. Both developmental stages and sexes moved considerable distances, were cap\able of crossing habitat boundaries, and showed successful inter-patch dispersal through the agricultural matrix.
3. The results suggest that P. griseoaptera exhibits movement behaviours that compensate for its inability to fly. It was concluded that the good dispersal ability exhibited by animals of both sexes and developmental stage is an important factor allowing P. griseoaptera to persist in fragmented agricultural landscapes.     

Completeness of Follow-Up Determines Validity of Study Findings: Results of a Prospective Repeated Measures Cohort Study

Background

Current reporting guidelines do not call for standardised declaration of follow-up completeness, although study validity depends on the representativeness of measured outcomes. The Follow-Up Index (FUI) describes follow-up completeness at a given study end date as ratio between the investigated and the potential follow-up period. The association between FUI and the accuracy of survival-estimates was investigated.

Methods

FUI and Kaplan-Meier estimates were calculated twice for 1207 consecutive patients undergoing aortic repair during an 11-year period: in a scenario A the population’s clinical routine follow-up data (available from a prospective registry) was analysed conventionally. For the control scenario B, an independent survey was completed at the predefined study end. To determine the relation between FUI and the accuracy of study findings, discrepancies between scenarios regarding FUI, follow-up duration and cumulative survival-estimates were evaluated using multivariate analyses.

Results

Scenario A noted 89 deaths (7.4%) during a mean considered follow-up of 30±28months. Scenario B, although analysing the same study period, detected 304 deaths (25.2%, P<0.001) as it scrutinized the complete follow-up period (49±32months). FUI (0.57±0.35 versus 1.00±0, P<0.001) and cumulative survival estimates (78.7% versus 50.7%, P<0.001) differed significantly between scenarios, suggesting that incomplete follow-up information led to underestimation of mortality. Degree of follow-up completeness (i.e. FUI-quartiles and FUI-intervals) correlated directly with accuracy of study findings: underestimation of long-term mortality increased almost linearly by 30% with every 0.1 drop in FUI (adjusted HR 1.30; 95%-CI 1.24;1.36, P<0.001).

Conclusion

Follow-up completeness is a pre-requisite for reliable outcome assessment and should be declared systematically. FUI represents a simple measure suited as reporting standard. Evidence lacking such information must be challenged as potentially flawed by selection bias.     

Cytogeography of Solidago gigantea (Asteraceae) and its invasive ploidy level

Aim Newly formed polyploids experience problems of establishment and spread similar to those faced by newly introduced alien species. To understand the significance of polyploidy in biological invasions, we mapped the distribution of ploidy levels in Solidago gigantea Aiton in its native range in North America, and in Europe and East Asia where it is invasive. Location North America, Europe and East Asia. Methods Flow cytometry was used to measure ploidy levels in a total of 834 plants from 149 locations. Together with data from contributors and a literature review, ploidy‐level data were assembled for 336 locations. Cytogeographical maps from North America and Europe were prepared, incorporating new and previously published ploidy‐level data. Results In the native range, diploid, tetraploid and hexaploid plants were found, and also one triploid and one pentaploid plant (2n = 3x and 2n = 5x, respectively, each being new reports for this species). There was a high degree of geographical separation among the ploidy levels, and populations with mixed ploidy were rare. However, four zones were identified where plants of different ploidy could come into contact. In Europe and East Asia, only tetraploid plants were found. Main conclusions The geographical pattern in North America suggests that the ploidy levels are ecologically differentiated, although further investigations are needed to identify the nature of these differences. Alien populations appear to be exclusively tetraploid, but it is not clear whether this is because tetraploids were selectively introduced or because diploids were unsuccessful. In any case, comparisons between native and introduced populations need to account for ploidy level.