Incorporation of simvastatin in PLLA membranes for guided bone regeneration: effect of thermal treatment on simvastatin release

Electrospun membranes based on biodegradable polymers are promising materials to be used for guided bone regeneration (GBR) therapy. The incorporation of osteostimulatory compounds can improve the biofunctionality of those membranes, making them active players in bone regeneration. Simvastatin has been shown to promote osteogenic differentiation both in vitro and in vivo. However, in most of these systems, the drug was quickly released, not matching the pace of bone regeneration. The aim of this study was to develop poly(l-lactic acid) (PLLA) membranes containing simvastatin (SV) that have a prolonged drug release rate, compatible with GBR applications. To this end, SV was mixed with PLLA and electrospun. The membranes were subjected to a thermal treatment in order to increase the crystallinity of PLLA. Morphological, structural and chemical properties of the electrospun membranes were characterized. The effect of the thermal treatment on the release profile of SV was evaluated by near physiological release experiments at 37 °C. The osteostimulatory potential was determined by in vitro culture of the membranes with rat bone marrow stromal cells (rBMSCs). The results confirmed that the thermal treatment led to an increase in polymer crystallinity and a more sustained release of SV. In vitro assays demonstrate cellular proliferation over time for all the membranes and a significant increase in osteogenic differentiation for the membranes containing SV subjected to thermal treatment.

Thermal treatment resulted in a sustained release of simvastatin and a positive response from rBMSCs.     

Oxygenation Status in Chronic Leg Ulcer After Topical Hemoglobin Application May Act as a Surrogate Marker to Find the Best Treatment Strategy and to Avoid Ineffective Conservative Long-term Therapy

Purpose

Chronic leg ulcers can be a challenge to treat and long-term therapy a significant cost factor in western public health budgets. Objective wound assessment assays enabling selection of appropriate wound therapy regimes would be desirable. Oxygenation status in ulcer tissue has obtained increased attention as a relevant factor in wound healing. To increase oxygenation in wounds, a topical hemoglobin spray was developed. Although favorable results have been noted, the link between clinical efficacy and the mode of action has not been demonstrated. The aims were to determine if changes in tissue oxygenation can be measured after topical application of hemoglobin on chronic wounds and to evaluate the findings in terms of therapy strategies.

Procedures

Photoacoustic imaging was used to measure the local oxygen saturation (StO₂) in leg ulcers before and after hemoglobin spray treatment. Sclerosis of the leg ulcers was histopathologically graded and the change in wound size was documented in a follow-up examination.

Results

Measuring 49 patients, an increase in StO₂ after topical hemoglobin application from on average 66.1 to 71 % (p = 0.017) after 20 min was observed. Depending on the increase in StO₂ (>10 % or <10 %) patients were stratified into a Responder and a Non-Responder group. Wound size significantly decreased in the Responder Group (p = 0.001), while no significant difference in the Non-Responder group (p = 0.950) was noted.

Conclusion

Our findings suggest that the likelihood of wound healing under conservative therapy can be predicted by measuring changes in StO₂ after topical hemoglobin application. This assay may reduce treatment time and costs by avoiding ineffective conservative long-term therapy.

Trial Registration

German Clinical Trials Register: DRKS00005993

     

The impact of the composition of the bone marrow graft on engraftment and graft-versus-host disease

To study the impact of the composition of the bone-marrow graft on engraftment and graft-versus-host disease (GvHD), we analyzed the data on 29 patients with acute leukemia in remission and 11 patients with aplastic anemia. All of them received bone-marrow grafts from HLA matched, MLC nonreactive, sibling donors, were nursed in laminar down-flow isolators with selective gut decontamination, and received GvHD prophylaxis with methotrexate. The number of nucleated cells in the marrow graft/kg body weight of the recipient had no relation with the rapidity of engraftment or with the occurrence and severity of GvHD. The number of hematopoietic progenitor cells (CFUc)/kg had a weak, but significant, correlation with both the number of neutrophils at day 30 post BMT and with the day at which the reticulocytes passed the 10% level. The number of T cells/kg did not show any correlation with either the occurrence or the severity of GvHD. Our data show that the concentration of hematopoietic progenitor cells in the graft correlates with the rapidity of engraftment. However, within the range of numbers of T cells infused in this study, no correlation is present between T cells in the graft and GvHD. Therefore, nearly complete depletion of marrow grafts of T cells is probably necessary to effectively decrease the incidence of GvHD.     

Down-regulation of hepatic lipase expression by elevation of cAMP in human hepatoma but not adrenocortical cells

Expression of hepatic lipase (HL) in the liver is reduced during prolonged fasting. This effect is mainly mediated via catecholamines, which signal through elevation of Ca(i)(2+) as well as cAMP. We have studied the effect of cAMP on HL expression in cell culture. Overnight incubation of HepG2 cells with 10-300microM 8-bromo-cyclic AMP resulted in a dose-dependent, up to 50% reduction in secretion of HL, but had no effect on secretion of alpha(1)-antitrypsin or overall protein synthesis. HL mRNA levels were decreased 1.5 fold, as determined by semi-quantitative and real-time RT-PCR. In HepG2 cells transiently transfected with human HL (-685/+13) or rat HL (-446/+9) promoter-reporter constructs, cAMP induced a similar dose-dependent suppression of HL promoter activity. cAMP responsiveness in HepG2 cells was mediated by a conserved 10-bp response element at -45/-36, that represents a potential binding site for CCAAT/enhancer-binding protein beta (C/EBPbeta). cAMP reduced expression of the 45kDa C/EBPbeta protein and binding of C/EBPbeta to the proximal promoter region of the human HL gene by 50%, as determined by immunoblotting and chromatin immunoprecipitation assay, respectively. In human H295R adrenocortical cells, cAMP failed to suppress HL promoter activity, and only slightly reduced C/EBPbeta expression. We conclude that the fall in HL expression during prolonged fasting may be mediated through elevation of cAMP and lowering of C/EBPbeta expression.     

Real-Time Genomic Epidemiological Evaluation of Human Campylobacter Isolates by Use of Whole-Genome Multilocus Sequence Typing

Sequence-based typing is essential for understanding the epidemiology of Campylobacter infections, a major worldwide cause of bacterial gastroenteritis. We demonstrate the practical and rapid exploitation of whole-genome sequencing to provide routine definitive characterization of Campylobacter jejuni and Campylobacter coli for clinical and public health purposes. Short-read data from 384 Campylobacter clinical isolates collected over 4 months in Oxford, United Kingdom, were assembled de novo. Contigs were deposited at the pubMLST.org/campylobacter website and automatically annotated for 1,667 loci. Typing and phylogenetic information was extracted and comparative analyses were performed for various subsets of loci, up to the level of the whole genome, using the Genome Comparator and Neighbor-net algorithms. The assembled sequences (for 379 isolates) were diverse and resembled collections from previous studies of human campylobacteriosis. Small subsets of very closely related isolates originated mainly from repeated sampling from the same patients and, in one case, likely laboratory contamination. Much of the within-patient variation occurred in phase-variable genes. Clinically and epidemiologically informative data can be extracted from whole-genome sequence data in real time with straightforward, publicly available tools. These analyses are highly scalable, are transparent, do not require closely related genome reference sequences, and provide improved resolution (i) among Campylobacter clonal complexes and (ii) between very closely related isolates. Additionally, these analyses rapidly differentiated unrelated isolates, allowing the detection of single-strain clusters. The approach is widely applicable to analyses of human bacterial pathogens in real time in clinical laboratories, with little specialist training required.     

Longitudinal associations in adolescence between cortisol and persistent aggressive or rule-breaking behavior

Although several studies have associated antisocial behavior with decreased cortisol awakening responses (CAR), studies in adolescent samples yielded inconsistent results. In adolescence however, the CAR develops and antisocial behavior is heterogeneous in type and persistence. Therefore this longitudinal study compared persistent aggressive and rule-breaking adolescents to low aggressive and rule-breaking adolescents on the development of the CAR from ages 15 to 17 (N = 390). Persistently high aggressive adolescents showed decreased cortisol levels at awakening consistently over the years (Δχ²(1) = 6.655, p = .01) as compared to low aggressive adolescents. No differences between adolescents showing persistent high rule-breaking and low rule-breaking were found. This longitudinal study is the first to show that persistent aggression, but not rule-breaking behavior, is related to neurobiological alterations. Moreover, despite development of the CAR over adolescence, the decrease in cortisol is consistent over time in persistent high aggressive adolescents, which is an important prerequisite for the prediction of persistent aggression.