Spatiotemporal presentation of exogenous SDF-1 with PLGA nanoparticles modulates SDF-1/CXCR4 signaling axis in the rodent cortex.

Stromal cell-derived factor-1 (SDF-1) and its key receptor CXCR4 have been implicated in directing cellular recruitment for several pathological/disease conditions thus also gained considerable attention for regenerative medicine. One regenerative approach includes sustained release of SDF-1 to stimulate prolonged stem cell recruitment. However, the impact of SDF-1 sustained release on the endogenous SDF-1/CXCR4 signaling axis is largely unknown as auto-regulatory mechanisms typically dictate cytokine/receptor signaling. We hypothesize that spatiotemporal presentation of exogenous SDF-1 is a key factor in achieving long-term manipulation of endogenous SDF-1/CXCR4 signaling. Here in the present study, we sought to probe our hypothesis using a transgenic mouse model to contrast the spatial activation of endogenous SDF-1 and CXCR4 in response to exogenous SDF-1 injected in bolus or controlled release (PLGA nanoparticles) form in the adult rodent cortex. Our data suggests that the manner of SDF-1 presentation significantly affected initial CXCR4 cellular activation/recruitment despite having similar protein payloads over the first 24 h (∼30 ng for both bolus and sustained release groups). Yet, one week post-injection, this response was negligible. Therefore, the transient nature CXCR4 recruitment/activation in response to bolus or controlled release SDF-1 indicated that cytokine/receptor auto-regulatory mechanisms may demand more complex release profiles (i.e. delayed and/or pulsed release) to achieve sustained cellular response.

Tunable delayed controlled release profile from layered polymeric microparticles.

Composite microparticles (MPs) with layered architecture, engineered from poly(L-lactic acid) (PLLA) and poly(D,L-lactic-co-glycolic acid) (PLGA), are promising devices for achieving the delayed release of proteins. Here, we build on a water-in-oil-in-oil-in-water emulsion method of fabricating layered MPs with an emphasis on modulating the delay period of the protein release profile. Particle hardening parameters (i.e. polymer precipitation rate and total hardening time) following water-in-oil-in-oil-in-water emulsions are known to affect MP structure such as the core/shell material and cargo localization. We demonstrate that layered MPs fabricated with two different solvent evaporation parameters not only alter polymer and protein distribution within the hardened MPs, but also affect their protein release profiles. Secondly, we hypothesize that ethanol (EtOH), a semi-polar solvent miscible in both the solvent (dichloromethane; DCM) and non-solvent aqueous phases, likely alters DCM and water flux from the dispersed oil phase. The results reveal that EtOH affects protein distribution within MPs, and may also influence MP structural properties such as porosity and polymer distribution. To our knowledge, we are the first to demonstrate EtOH as a means for modulating critical release parameters from protein-loaded, layered PLGA/PLLA MPs. Throughout all the groups in the study, we achieved differential delay periods (between 0 - 30 days after an initial burst release) and total protein release periods (~30 - >58 days) as a function of solvent evaporation parameters and EtOH content. The layered MPs proposed in the study potentially have wide-reaching applications in tissue engineering for delayed and sequential protein release.

A comparative study using thrombin generation and three different INR methods in patients on Vitamin K antagonist treatment.

INTRODUCTION: Vitamin K antagonist (VKA) treatment requires routine monitoring using the international normalized ratio (INR). However, different INR assays may vary in their results. The aim of this study was to assess the agreement of three different INR methods, compared with thrombin generation, in patients on VKA treatment. METHODS: Sixty patients attending the Anticoagulation Clinic at Mater Dei Hospital (Msida, Malta) for VKA monitoring between August and September 2015 were enrolled. The INR was tested using a point-of-care (POC) device (CoaguChek XS Plus, Roche Diagnostics) for both capillary and venous blood samples, a photo-optical (Sysmex CS-2100i/CA-1500, Siemens) and a mechanical clot detection system (Thrombolyzer XRC, Behnk Elektronik). All assays used human recombinant thromboplastin as reagent. Thrombin generation was performed using the calibrated automated thrombogram. RESULTS: There was a negative curvilinear correlation between the endogenous thrombin potential and different INR assays (r≤-.75) and a strong positive linear correlation between the CoaguChek XS Plus on capillary samples and the other INR methodologies (r≥.96). CONCLUSION: All different INR assays showed good correlation with the thrombin generation potential. The POC INR showed one of the highest correlation coefficients with thrombin generation, confirming the POC devices as an accurate, valid alternative to laboratory INR in VKA patients.

Evaluating the benefits of vaccination when used in combination with stamping-out measures against hypothetical introductions of foot-and-mouth disease into New Zealand: a simulation study.

AIMS: To evaluate the benefits of vaccination against simulated outbreaks of foot-and-mouth disease (FMD) in New Zealand, when applied as an additional measure to stamping-out. METHODS: A simulation modelling approach was used. The study population comprised all known farms in New Zealand with FMD-susceptible livestock. Infection was seeded into three different areas of New Zealand. Transmission mechanisms included direct and indirect contacts, local spread and airborne spread. Efficacies of some of the stamping-out measures were varied. Vaccination strategies involved different start times, size and type of vaccination zone, and species vaccinated. Personnel resources for vaccination were varied as was the herd immunity profile following vaccination. Altogether, 336 models were specified, with 100 iterations conducted for each model. Generalised linear modelling and boosted regression trees were used to evaluate which variables had the biggest effect on the number of infected premises (IP), epidemic duration and area under control. RESULTS: Vaccination, when used as an adjunct to the standard stamping-out programme, significantly reduced the outbreak size. Vaccination reduced the median number of IP by 26 (95% CI=18-35), epidemic duration by 16 (95% CI=13-19) days and area under control by 474 (95% CI=250-699) km2 when there was no airborne spread; and when there was airborne spread the median reduction was 87 (95% CI=70-105) IP, 32 (95% CI=28-35) days and 898 (95% CI=665-1139) km2, respectively. Multivariable analyses showed that starting vaccination 11 days after first detection of FMD produced greater benefits than starting 16 or 21 days after detection. Increasing vaccination zones resulted in increased benefits. Boosted regression tree analyses showed that the most influential variables on the outcome measures were interval to first detection, incursion location, whether there was airborne spread or not and herd immunity profile. CONCLUSIONS AND CLINICAL RELEVANCE: This study showed that there are benefits to the use of vaccination in combination with a stamping-out policy for control of FMD outbreaks under New Zealand conditions. The optimal vaccination strategy was identified as being a 3-5 km radius suppressive vaccination zone deployed between 11-16 days after first detection. Vaccination had a greater benefit during larger outbreaks, such as when there was airborne transmission. The key factors which were identified from this study will help inform New Zealand's competent authority on how best to deploy vaccination to further strengthen its approach to FMD eradication should New Zealand ever experience an outbreak.

The use and success of cold coagulation for the treatment of high grade squamous cervical intra-epithelial neoplasia: a retrospective review.

OBJECTIVE: Cold coagulation is recognised as a viable, cost-effective and successful treatment for cervical intraepithelial neoplasia (CIN), being used less frequently than excisional treatments for high grade lesions. We set out to demonstrate successful long term follow-up of patient with high grade CIN treated with cold coagulation. STUDY DESIGN: We conducted a retrospective review over a one-year period of women with biopsy-proven CIN 2 and 3 who were treated with cold coagulation to the cervix, attending the colposcopy service of a large tertiary referral hospital. We examined follow-up cervical smear data for three years post treatment of low and high grade CIN, evaluated the success of treatment and re-treatment rates. RESULTS: 93 patients were included in our study, with 39 (41.9%) having CIN 1 and 54 (58.1%) diagnosed with CIN 2 or 3. Follow-up smears revealed low levels of recurrent high grade changes in both groups, with 31 (79.5%) of our CIN 1 group having a negative smear one year following treatment with cold coagulation, compared to 44 (81.1%) of patients with CIN 2 and 3. Successful primary treatment (i.e. no requirement for further treatment after 3 year follow-up) occurred in 33 (84.6%) of the CIN 1 group, and 42 (77.7%) of the CIN 2/3 group, demonstrating no statistical significance between re-treatment rates between both groups. CONCLUSIONS: This study demonstrates the effectiveness of cold coagulation for the treatment of high grade cervical intraepithelial neoplasia. High success rates, and low re-treatment rates confirm that this is an acceptable primary treatment for CIN 2 and 3.

Relationships between range access as monitored by radio frequency identification technology, fearfulness, and plumage damage in free-range laying hens.

Severe feather-pecking (SFP), a particularly injurious behaviour in laying hens (Gallus gallus domesticus), is thought to be negatively correlated with range use in free-range systems. In turn, range use is thought to be inversely associated with fearfulness, where fearful birds may be less likely to venture outside. However, very few experiments have investigated the proposed association between range use and fearfulness. This experiment investigated associations between range use (time spent outside), fearfulness, plumage damage, and BW. Two pens of 50 ISA Brown laying hens (n=100) were fitted with radio frequency identification (RFID) transponders (contained within silicone leg rings) at 26 weeks of age. Data were then collected over 13 days. A total of 95% of birds accessed the outdoor run more than once per day. Birds spent an average duration of 6.1 h outside each day over 11 visits per bird per day (51.5 min per visit). The top 15 and bottom 15 range users (n=30), as determined by the total time spent on the range over 13 days, were selected for study. These birds were tonic immobility (TI) tested at the end of the trial and were feather-scored and weighed after TI testing. Birds with longer TI durations spent less time outside (P=0.01). Plumage damage was not associated with range use (P=0.68). The small group sizes used in this experiment may have been conducive to the high numbers of birds utilising the outdoor range area. The RFID technology collected a large amount of data on range access in the tagged birds, and provides a potential means for quantitatively assessing range access in laying hens. The present findings indicate a negative association between fearfulness and range use. However, the proposed negative association between plumage damage and range use was not supported. The relationships between range use, fearfulness, and SFP warrant further research.

Theileria orientalis: a review.

Theileria orientalis (also known historically as T. sergenti and T. buffeli) is responsible for benign or non-transforming theileriosis, and exerts its major effect through erythrocyte destruction. The life cycle of T. orientalis is essentially similar to that of other Theileria species, except that the schizonts do not induce transformation and fatal lymphoproliferation. The pathogenesis of anaemia as a result of infection is not clearly established and may be multifaceted. Clinical signs of weakness, reluctance to walk and abortion are early but non-specific indications of disease, particularly if accompanied by a history of cattle being moved. Physical examination may reveal pallor (pale eyes, vaginal mucosa), pyrexia, and elevated heart and respiratory rates. T. orientalis is an economically important parasite of cattle in New Zealand, Australia and Japan, especially where naïve animals are introduced into an endemic area or in animals under stress. Increased awareness of the risks posed by the parasite is required to enable management practices to be implemented to minimise its impact.

Stress and dietary factors modify boar sperm for processing.

This paper reviews stresses boar sperm undergo during processing and presents preliminary results of dietary modification that minimize this damage. Processing for artificial insemination (AI) stresses boar sperm by osmotic effects; altering cell size, shape and membranes; intracellular ice formation; and production of reactive oxygen species (ROS). Sperm response to ROS is concentration-dependent, with low levels activating the ERK pathway to stimulate tyrosine phosphorylation (Tyr-P) and capacitation, but high concentrations or inappropriately timed onset of ROS pathways can harm sperm. Fresh boar sperm exposed to ROS increased intracellular hydrogen peroxide (H(2) O(2) ) phospholipase and lipid peroxidation, maintained viability but lost motility and underwent acrosome reactions (AR). Direct incorporation of lipids ± the antioxidant Vitamin E improves the survival of liquid- and frozen-stored semen. Boars fed dietary flaxseed for 8 weeks to increase n-3 fatty acids displayed improved sperm morphology (p < 0.05), increased membrane fluidity (p < 0.05) and better retention of motility and viability during 5-7 day storage (p < 0.05). Processes reducing oxidative damage to stored sperm should be evaluated.

Clinical multicenter evaluation of a new FXa-based Antithrombin assay.

INTRODUCTION: The determination of functional Antithrombin is a central part of thrombophilia screening. In this multicenter study, a new FXa-based method (INNOVANCE® Antithrombin) was evaluated on four different analyzers. METHODS: The INNOVANCE Antithrombin method was evaluated by precision and reference interval studies and by comparing the new method with established methods through parallel measurement of samples from 249 patients and 151 apparently healthy individuals. RESULTS: The INNOVANCE Antithrombin assay demonstrated on all analyzers repeatability coefficients of variation (CVs) ≤ 3.2% and within-device and between-run CVs ≤ 6.9%. The reference intervals of all analyzers are comparable with 2.5th percentiles between 80% and 85% of normal. The INNOVANCE Antithrombin and the FIIa-based Berichrom® AT III (A) methods demonstrated good concordance with correlation coefficients of r = 0.908 or higher. The INNOVANCE Antithrombin method demonstrated furthermore an excellent comparability with the STA® Antithrombin III assay and an acceptable comparability with the Coamatic® LR Antithrombin assay. The patients with congenital deficiency (n = 31) were identified with all assays except for the patients carrying the P41L heparin-binding site mutation, which was only identified with the INNOVANCE Antithrombin and the STA Antithrombin III methods. CONCLUSION: The INNOVANCE Antithrombin assay has high sensitivity for Antithrombin deficiencies and is reliable, precise and suitable for routine clinical use.

Evaluation of a new venom-based clotting assay of protein C.

Congenital protein C deficiency significantly increases the risk of venous thromboembolism, a serious and potentially lethal condition. Protein C levels can be determined by chromogenic, clotting and antigenic assays, each type of assay has differences in specificity and sensitivity to protein C deficiency. In principle, clotting-based assays of protein C are preferred over chromogenic assays, as they can detect some rare mutations that are missed by the chromogenic assay, however, clotting-based assays may be prone to inaccuracy because of poor specificity. We have evaluated a new venom-based clotting assay of protein C, and optimized it for use on Sysmex CA-1500 analyser. The assay was linear from 0 to 130 U/dl, a normal plasma demonstrated good inter-assay precision, with a coefficient of variation of 4.8%. The assay compared well with antigenic- and venom-based chromogenic protein C assay in normal individuals, subjects with lupus anticoagulant, and subjects with FV Leiden. Median protein C levels by clotting, chromogenic and antigen for the three subject groups were 108 U/dl, 108 IU/dl and 109 IU/dl for normal subjects, 94 U/dl, 106 IU/dl and 103 IU/dl for subjects with lupus anticoagulant, and 102 U/dl, 104 IU/dl and 100 IU/dl for subjects heterozygous for FV Leiden. Comparing levels of clotting protein C with protein C antigen by ratio (clotting/antigen), the three groups showed small differences that did not quite reach statistical significance, (mean ratios ranged from 0.95 to 1.01, anovaP = 0.0561), the lowest ratio was with the lupus anticoagulant group. Comparing clotting assay with chromogenic assay by ratio (clotting/chromogenic), the three groups did show a statistically significant difference (P = 0.0033) which was due to a difference in mean ratios between normal and lupus anticoagulant groups (ratios 1.00 and 0.91, respectively, P < 0.01). There was no statistical difference in any of the groups when comparing chromogenic protein C with protein C antigen (mean ratios ranged from 1.02 to 1.05, P = 0.3925). In a normal sample, the clotting-based protein C level was unaffected by increasing FVIII level by up to 1000 IU/dl, using intermediate purity FVIII concentrate. The new assay is considered to be a suitable assay for the routine diagnosis of protein C deficiency.

Correlation of serum CA125 with stage, grade and survival of patients with epithelial ovarian cancer at a single centre.

To evaluate the relationship between serum CA125 tumour marker level before and after surgery of epithelial ovarian carcinoma and assess its potential role as a prognostic factor. A retrospective review of 87 patients with epithelial ovarian carcinoma at a single centre between January 2001 and December 2005 was performed. Serum CA125 levels were assessed for their relationship to pathological stage, tumour grade, tumour volume and age as well as overall survival. A total of 75 patients, mean age 58.94 years and median follow-up of 24 months were included in the analysis. While the preoperative CA125 level did not correlate significantly with stage, tumour grade or survival, the postoperative CA125 correlated to FIGO stage (p<0.0001), tumour grade (p<0.0001) and overall survival (p=0.01). Reduced survival was noted with increasing age at the time of surgery (p=0.009) and bulk of the residual disease postoperatively (p=0.011).

Modeling anaerobic digestion of dairy manure using the IWA Anaerobic Digestion Model no. 1 (ADM1).

The Anaerobic Digestion Model No. 1 (ADM1) can be used to describe treatment of dairy manure once manure characteristics have been incorporated in the model. In this paper a parameter set is presented that can be used with ADM1 for simulation of dairy manure digester performance. Model results have been verified with bench-scale experiments and reported data from full-scale systems. Model predictions fit experimental data best for biogas composition and digester effluent COD. Simulated biogas productions were inconsistent with measurements from three different digesters. The model overpredicted acetogenesis, resulting in higher simulated than observed acetate concentrations. However, total volatile acid concentrations were simulated reasonably well. The model consistently predicted higher inorganic nitrogen than measured or reported results, indicating a need for further research in that area. The presented model and associated parameter set can be used to simulate and optimize the performance of full-scale dairy manure digesters.

Introducing laparoscopic management of ectopic pregnancy to a busy gynaecology unit--is it worthwhile?

A review was carried out on 26 consecutive women undergoing surgery for ectopic pregnancy in the Limerick Regional Hospital, Ireland from April 2000 to April 2002. 13 were managed laparoscopically and 13 had laparotomy. There were no significant differences in age, parity or gestational age. 3 patients had previous ectopic pregnancy. 12/13 from the laparotomy group had a diagnostic laparoscopy prior to laparotomy. Anaesthetic time differed by 21.2 minutes with laparotomy being done faster than the laparoscopy group while operative time was 7.3 minutes longer in the laparoscopic group. The laparoscopic approach was associated with lower intraoperative blood loss (<50 ml vs 413.1 ml), less post operative analgesia requirement, shorter hospital stay (2.4 days vs 4.5 days), faster return to work (2 weeks vs 4 weeks) and less subsequent wound infection. Operative laparoscopy also has the advantage of being a diagnostic as well as a therapeutic tool in one procedure.

Does surgical staging of clinical Stage I endometrial carcinoma significantly alter adjuvant management?

Full surgical staging for endometrial cancer has been advocated since 1988, when FIGO redefined stage of disease on the basis of surgico-pathological criteria. However, such extensive surgery, performed on often elderly women, must counterbalance any increased morbidity against the potential gains of altered subsequent management. We report a 2-year review of 22 patients undergoing full surgical staging for Stage I endometrial cancer. Nine women (41%) had altered adjuvant treatment as a result. In seven patients, decisions regarding postoperative radiotherapy treatment were influenced and two women had an unsuspected ovarian tumour. Postoperative complications were minor, apart from one wound dehiscence. There were no perioperative deaths and no blood transfusions. Surgical staging was found to be a safe intervention with a significant impact on adjuvant management.