Recombinant mouse granulocyte chemotactic protein-2: production in bacteria, characterization, and systemic effects on leukocytes.

Granulocyte chemotactic protein-2 (GCP-2) is an important neutrophil chemotactic factor in the mouse that belongs to the CXC chemokine family. Although the local tissular effects of chemokines are well known, only recently has the systemic regulation of leukocytes become accepted. To study the pharmacokinetics of mouse GCP-2 and the systemic effects on leukocytes, we expressed a potent natural isoform of mouse GCP-2, GCP-2(9-78), in Escherichia coli and produced electrophoretically pure material. GCP-2(9-78) was 10-fold more potent to chemoattract neutrophils than recombinant GCP-2(5-78). After intravenous (i.v.) injection in mice, GCP-2(9-78) persisted in the circulation with an average half-life of 42 min. When a bolus of 1 mg/kg recombinant mouse GCP-2(9-78) was injected systemically, a significant effect on circulating leukocytes was observed. After a neutropenic phase, at its height at 1 h after injection, neutrophil numbers increased to a maximum at 4 h postinjection, and a concomitant decrease in lymphocyte numbers was observed. In control mice injected with isotonic saline, changes in leukocyte numbers were less pronounced and followed a different kinetic. Whereas tissular neutrophil chemotaxis to GCP-2 is influenced by gelatinase B, the systemic effects on neutrophilia and lymphopenia were not different in gelatinase B-deficient and wild-type mice. These data reinforce the idea that chemokines, including GCP-2, influence the homeostasis of circulating leukocyte numbers.     

Identification,molecular cloning,and characterization of the chromosome 12 breakpoint cluster region of uterine leiomyomas

Recent molecular cytogenetic analysis of uterine leiomyoma cell lines with chromosome 12 aberrations has indicated that their chromosome 12 breakpoints map between linkage locus D12S8 and the CHOP gene. Here, we report fluorescence in situ hybridization (FISH) and molecular cloning studies of the chromosome 12 breakpoints in a panel of seven such uterine leiomyoma cell lines; five with the frequently observed t(12; 14)(q15;q24), one with t(12;15)(q15;q24), and one with ins(12;11)(q14;q21 qter). Directional chromosome walking studies starting from D12S8 and the CHOP gene resulted in the isolation of a relatively large number of overlapping YAC clones, including Y5355 (465 kbp), Y7673 (360 kbp), and Y9899 (275 kbp). In total, the inserts of these three YAC clones span an 800 kbp long and presumably contiguous stretch of human genomic DNA. All chromosome 12 breakpoints of the uterine leiomyoma cell lines studied were found by FISH analysis to be mapping within a 445 kbp subfragment of this region and, furthermore, to be dispersed over a DNA region which is at least 150 kbp in size. The chromosome 12 breakpoint of t(12;14)(q15;q24) in uterine leiomyoma cell line LM-30.1/SV40 was tentatively mapped within the 60 kbp region between YAC clones Y9899 and Y5355. From this 60 kbp region and close to sequencetagged site RM99, we isolated probe pRM 118-A, which showed in Southern blot analysis that it detected a rearrangement in LM-30.1/SV40 DNA, and generated restriction maps of the normal and rearranged genomic DNA regions detected with this probe. Finally, we molecularly cloned part of one of those rearranged DNA fragments using a vectorette-PCR-based technique and demonstrated that it consisted of 12q13-q15 sequences fused to DNA sequences derived from 14q23-24 and most likely represented the translocation junction on der(14) in LM-30.1/SV40 cells. Our studies strongly suggest that we have identified and isolated the uterine leiomyoma cluster region of chromosome 12 breakpoints, which we designate ULCR12, and molecularly cloned and characterized the der(14) translocation junction in cells derived from a uterine leiomyoma carrying the frequently observed t(12;14)(q15;q24).     

Reporting the results of meta-analyses: a plea for incorporating clinical relevance referring to an example

Background Context

The results of meta-analyses are frequently reported, but understanding and interpreting them is difficult for both clinicians and patients. Statistical significances are presented without referring to values that imply clinical relevance.

Formulation of Cell-Based Medicinal Products: A Question of Life or Death?

The formulation of cell-based medicinal products (CBMPs) poses major challenges because of their complexity, heterogeneity, interaction with their environment (e.g., the formulation buffer, interfaces), and susceptibility to degradation. These challenges can be quality, safety, and efficacy related. In this commentary we discuss the current status in formulation strategies of off-the-shelf and non-off-the-shelf (patient-specific) CBMPs and highlight advantages and disadvantages of each strategy. Analytical tools for the characterization and stability assessment of CBMP formulations are addressed as well. Finally, we discuss unmet needs and make some recommendations regarding the formulation of CBMPs.     

Initial steps towards an evidence-based classification system for golfers with a physical impairment

Purpose: The present narrative review aims to make a first step towards an evidence-based classification system in handigolf following the International Paralympic Committee (IPC). It intends to create a conceptual framework of classification for handigolf and an agenda for future research. Method: Pubmed was searched on three themes: “Classification in Paralympic sports”, “Performance determining factors in golf” and “Impact of impairments on golf performance”. IPC-regulations were gathered on the IPC-website and their official publications. Results: In developing a classification system conform IPC-regulations, the main challenge is to identify the activity limitation caused by the impairment, not influenced by training, talent or motivation. Timing, accuracy and control, work per joint, range of motion, balance and flexibility are important performance determining factors in abled-bodied golf and should be considered when determining activity limitations in handigolf. Only five articles on handigolf were found, mainly addressing the asymmetric golf movement. Based on the present review, a conceptual framework for classification was developed, while a future research agenda was designated. The conceptual framework presents factors that are essential for sports performance categorized under “technology”, “interface” and “athlete characteristics”. It also includes impairment related factors essential for determining eligibility and classification. Ideally, measures to be used during classification need to be resistant against training, natural development of the athlete’s talent and motivational changes. Conclusions: The conceptual framework and a multidimensional scientific research agenda will support further development of the knowledge base required for an evidence-based classification in handigolf, including multi-level analysis of player statistics, experimental analyses of biomechanics and modeling studies.

• Implications for Rehabilitation

• The main challenge in developing an evidence-based classification system conform IPC-regulations is defining eligibility criteria and sport classes based on activity limitation caused by only the impairment and not affected by training, talent and motivation. It is expected that a transparent classification system, a lively competition and admission to the Paralympic program will further promote participation in disabled golf.

• Timing, accuracy and control, work per joint, range of motion, balance and flexibility are of greater importance for golf performance in able-bodied golfers and expected to be of interest to incorporate in classification for handigolf.

• Side and level of amputation influence activity limitation in the asymmetric golf movement, and should be incorporated in classification.

• The proposed conceptual framework is fundamental to the research agenda that must further generate the knowledge-base to determine activity limitations caused by different impairments in handigolf and may serve as a guideline for other Paralympic sports in the development of evidence-based classification.

     

Mesoporous Silica Nanoparticle-Coated Microneedle Arrays for Intradermal Antigen Delivery

Purpose

To develop a new intradermal antigen delivery system by coating microneedle arrays with lipid bilayer-coated, antigen-loaded mesoporous silica nanoparticles (LB-MSN-OVA).

Methods

Synthesis of MSNs with 10-nm pores was performed and the nanoparticles were loaded with the model antigen ovalbumin (OVA), and coated with a lipid bilayer (LB-MSN-OVA). The uptake of LB-MSN-OVA by bone marrow-derived dendritic cells (BDMCs) was studied by flow cytometry. The designed LB-MSN-OVA were coated onto pH-sensitive pyridine-modified microneedle arrays and the delivery of LB-MSN-OVA into ex vivo human skin was studied.

Results

The synthesized MSNs demonstrated efficient loading of OVA with a maximum loading capacity of about 34% and the lipid bilayer enhanced the colloidal stability of the MSNs. Uptake of OVA loaded in LB-MSN-OVA by BMDCs was higher than that of free OVA, suggesting effective targeting of LB-MSN-OVA to antigen-presenting cells. Microneedles were readily coated with LB-MSN-OVA at pH 5.8, yielding 1.5 μg of encapsulated OVA per microneedle array. Finally, as a result of the pyridine modification, LB-MSN-OVA were effectively released from the microneedles upon piercing the skin.

Conclusion

Microneedle arrays coated with LB-MSN-OVA were successfully developed and shown to be suitable for intradermal delivery of the encapsulated protein antigen.

     

Left ventricular solid body rotation in non‐compaction cardiomyopathy: A potential new objective and quantitative functional diagnostic criterion?

BACKGROUND: Left ventricular (LV) twist originates from the interaction between myocardial fibre helices that are formed during the formation of compact myocardium in the final stages of the development of myocardial architecture. Since non-compaction cardiomyopathy (NCCM) is probably caused by intrauterine arrest of this final stage, it may be anticipated that LV twist characteristics are altered in NCCM patients, beyond that seen in patients with impaired LV function and normal compaction. AIMS: The purpose of this study was to assess LV twist characteristics in NCCM patients compared to patients with non-ischaemic dilated cardiomyopathy (DCM) and normal subjects. METHODS AND RESULTS: The study population consisted of 10 patients with NCCM, 10 patients with DCM, and 10 healthy controls. LV twist was determined by speckle tracking echocardiography. In all controls and DCM patients, rotation was clockwise at the basal level and counterclockwise at the apical level. In contrast, in all NCCM patients the LV base and apex rotated in the same direction. CONCLUSIONS: These findings suggest that 'LV solid body rotation', with near absent LV twist, may be a new sensitive and specific, objective and quantitative, functional diagnostic criterion for NCCM.