Towards uterus tissue engineering: a comparative study of sheep uterus decellularisation.

Uterus tissue engineering may dismantle limitations in current uterus transplantation protocols. A uterine biomaterial populated with patient-derived cells could potentially serve as a graft to circumvent complicated surgery of live donors, immunosuppressive medication and rejection episodes. Repeated uterine bioengineering studies on rodents have shown promising results using decellularised scaffolds to restore fertility in a partially impaired uterus and now mandate experiments on larger and more human-like animal models. The aim of the presented studies was therefore to establish adequate protocols for scaffold generation and prepare for future in vivo sheep uterus bioengineering experiments. Three decellularisation protocols were developed using vascular perfusion through the uterine artery of whole sheep uteri obtained from slaughterhouse material. Decellularisation solutions used were based on 0.5% sodium dodecyl sulphate (Protocol 1) or 2% sodium deoxycholate (Protocol 2) or with a sequential perfusion of 2% sodium deoxycholate and 1% Triton X-100 (Protocol 3). The scaffolds were examined by histology, extracellular matrix quantification, evaluation of mechanical properties and the ability to support foetal sheep stem cells after recellularisation. We showed that a sheep uterus can successfully be decellularised while maintaining a high integrity of the extracellular components. Uteri perfused with sodium deoxycholate (Protocol 2) were the most favourable treatment in our study based on quantifications. However, all scaffolds supported stem cells for 2 weeks in vitro and showed no cytotoxicity signs. Cells continued to express markers for proliferation and maintained their undifferentiated phenotype. Hence, this study reports three valuable decellularisation protocols for future in vivo sheep uterus bioengineering experiments.

Dose tailoring of adjuvant chemotherapy for breast cancer based on hematologic toxicities: further results from the prospective PANTHER study with focus on obese patients.

Background: Adjuvant chemotherapy (ACT) for breast cancer improves relapse-free survival (BCRFS) and overall survival. Differences in terms of efficacy and toxicity could partly be explained by the significant interpatient variability in pharmacokinetics which cannot be captured by dosing according to body surface area. Consequently, tailored dosing was prospectively evaluated in the PANTHER trial. Patients and methods: PANTHER is a multicenter, open-label, randomized phase III trial which compared tailored, dose-dense (DD) epirubicin/cyclophosphamide (E/C) and tailored docetaxel (D) (tDD) with standard interval 5-fluorouracil/E/C and D. The primary end point was BCRFS and the primary efficacy analysis has been previously published. In this secondary analysis, we aimed to retrospectively explore the concept of dose tailoring. Our two hypotheses were that BCRFS would not vary depending on the cumulative administered epirubicin dose; and that dose tailoring would lead to appropriate dosing and improved outcomes for obese patients, who are known to have worse prognosis and increased toxicity after DD ACT. Results: Patients treated with tDD had similar BCRFS regardless of the cumulative epirubicin dose (P = 0.495), while obese patients in this group [body mass index (BMI) ≥30] had improved BCRFS compared with nonobese ones (BMI <30) [hazard ratio (HR) = 0.51, 95% confidence interval (CI) 0.30-0.89, P = 0.02]. Moreover, tDD was associated with improved BCRFS compared with standard treatment only in obese patients (HR = 0.49, 95% CI 0.26-0.90, P = 0.022) but not in nonobese ones (HR = 0.79, 95% CI 0.60-1.04, P = 0.089). The differences were not formally statistically significant (P for interaction 0.175). There were no differences in terms of toxicity across the epirubicin dose levels or the BMI groups. Conclusions: Dose tailoring is a feasible strategy that can potentially improve outcomes in obese patients without increasing toxicity and should be pursued in further clinical studies. ClinicalTrials.gov identifier: NCT00798070.

First clinical experience with levodopa/carbidopa microtablets in Parkinson's disease.

BACKGROUND: Levodopa is the most effective symptomatic treatment throughout the course of Parkinson's disease, but as the disease progresses, there may be a need for individualized, fine-tuned treatments. AIM: To evaluate individualized levodopa/carbidopa dosing using microtablets dispensed with a dose dispenser, with respect to efficacy and usability as perceived by patients. METHODS: Patient records and dose dispenser reports from patients previously or currently treated with microtablets and a dose dispenser were reviewed, and a patient questionnaire concerning effect and usability was sent to patients. RESULTS: Eleven patient records, four dose dispenser reports and nine survey responses were obtained. The treatment effect was considered to be improved by six of nine patients. One-third found their bradykinesia to be improved, and the non-troublesome dyskinesia was unchanged according to a majority of patients; however, some experienced the duration and magnitude of troublesome dyskinesia to be worse. The usability was generally rated as good. The four dose dispenser reports obtained showed 97(±5)% total adherence. CONCLUSIONS: The experienced effect of treatment can, for some patients, be improved by the use of microtablets, and the dose dispenser was considered user-friendly. Further studies with a larger study population and prospective design are needed to confirm the results.

The effect of a dentifrice containing Magnolia extract on established plaque and gingivitis in man: a six-month clinical study.

AIM: The aim was to evaluate the clinical effect of a dentifrice containing 0.3% Magnolia extract on dental plaque and gingivitis. MATERIAL AND METHODS: The trial was a 6-month double-blind, stratified, randomized and 2-armed parallel group study. Forty-six subjects in the test group brushed their teeth with a dentifrice containing 0.3% Magnolia extract and 48 subjects in the control group brushed with a placebo dentifrice. Plaque and gingivitis were assessed at baseline, 3 and 6 months. RESULTS: There was a significantly larger gingivitis reduction in the Magnolia group than in the control group (0.26 ± 0.11 versus 0.11 ± 0.12) (P < 0.001). There was a greater increase in the total number of healthy gingival units Gingival Index (GI score 0) in the Magnolia group than in the control group (149% versus 31%) and a larger reduction in inflamed gingival units (GI score 2/3) (60% versus 30%). Furthermore, at sites with similar amounts of plaque, less clinical signs of gingival inflammation were observed in the Magnolia group than in the control group. CONCLUSION: Six months' unsupervised use of a dentifrice containing 0.3% Magnolia extract resulted in significantly greater gingivitis reduction than a corresponding control dentifrice.

Individually tailored treatment with epirubicin and paclitaxel with or without capecitabine as first-line chemotherapy in metastatic breast cancer: a randomized multicenter trial.

Anthracyclines and taxanes are active cytotoxic drugs in the treatment of early metastatic breast cancer. It is yet unclear whether addition of capecitabine to the combination of these drugs improves the treatment outcome. Patients with advanced breast cancer were randomized to first-line chemotherapy with a combination of epirubicin (Farmorubicin(®)) and paclitaxel (Taxol(®)) alone (ET) or in combination with capecitabine (Xeloda(®), TEX). Starting doses for ET were epirubicin 75 mg/m(2) plus paclitaxel 175 mg/m(2), and for TEX epirubicin 75 mg/m(2), paclitaxel 155 mg/m(2), and capecitabine 825 mg/m(2) BID for 14 days. Subsequently, doses were tailored related to side effects. Primary endpoint was progression-free survival (PFS); secondary endpoints were overall survival (OS), time to treatment failure (TTF), objective response (OR), safety and quality of life (QoL). 287 patients were randomized, 143 to ET and 144 to TEX. Median PFS was 10.8 months for patients treated with ET, and 12.4 months for those treated with TEX (HR 0.84, 95% CI 0.65-1.07, P = 0.16); median OS was 26.0 months for women in the ET versus 29.7 months in the TEX arm (HR 0.84, 95% CI 0.63-1.11, P = 0.22). OR was achieved in 44.8% (ET) and 54.2% (TEX), respectively (χ(2) 3.66, P = 0.16). TTF was significantly longer for patients treated with TEX, 6.0 months, versus 5.2 months following ET (HR 0.73, 95% CI 0.58-0.93, P = 0.009). Severe hematological side effects related to epirubicin and paclitaxel were evenly distributed between the treatment arms, mucositis, diarrhea, and Hand-Foot syndrome were significantly more frequent in the TEX arm. Toxicity-adjusted treatment with ET and TEX showed similar efficacy in terms of PFS, OS, and OR. In this trial with limited power, the addition of capecitabine to epirubicin and paclitaxel as first-line treatment did not translate into clinically relevant improvement of the outcome.

Negative impact of rAAV2 mediated expression of SOCS3 on the regeneration of adult retinal ganglion cell axons.

Intravitreal injections of recombinant ciliary neurotrophic factor (rCNTF) protect adult rat retinal ganglion cells (RGCs) after injury and stimulate regeneration, an effect enhanced by co-injection with a cAMP analogue (CPT-cAMP). This effect is partly mediated by PKA and associated signaling pathways, but CPT-cAMP also moderates upregulation of suppressor of cytokine signaling (SOCS) pathways after rCNTF injection, which will also enhance the responsiveness of RGCs to this and perhaps other cytokines. We now report that intravitreal injections of CPT-cAMP do not potentiate RGC axonal regeneration when CNTF is expressed via an adeno-associated viral vector (rAAV2), and concomitantly we show that increases in retinal SOCS mRNA expression are less when CNTF is delivered using the vector. We also directly tested the impact of elevated SOCS3 expression on the survival and regeneration of injured adult RGCs by injecting a bicistronic rAAV2-SOCS3-GFP vector into the vitreous of eyes in rats with a peripheral nerve graft sutured onto the cut optic nerve. Overexpression of SOCS3 resulted in an overall reduction in axonal regrowth and almost complete regeneration failure of RGCs transduced with the rAAV2-SOCS3-GFP vector. Furthermore, rAAV2-mediated expression of SOCS3 abolished the normally neurotrophic effects elicited by intravitreal rCNTF injections. In summary, CNTF delivery to the retina using viral vectors may be more effective than bolus rCNTF injections because the gene therapy approach has a less pronounced effect on neuron-intrinsic SOCS repressor pathways. Our new gain of function data using rAAV2-SOCS3-GFP demonstrate the negative impact of enhanced SOCS3 expression on the regenerative potential of mature CNS neurons.

Cohort study of patients with adrenal lesions discovered incidentally.

BACKGROUND: This prospective cohort study investigated the incidence, clinical features and natural history of incidentally discovered adrenal mass lesions (adrenal incidentaloma, AI) in an unselected population undergoing radiological examination. METHODS: During an 18-month period, all patients with AI were reported prospectively from all 19 radiology departments in western Sweden. Inclusion criteria were: incidentally discovered adrenal enlargement or mass lesion in patients without extra-adrenal malignancy on detection. Clinical and biochemical evaluation was performed on inclusion and after 24 months. Computed tomography (CT) of the adrenals was scheduled at 4, 12 and 24 months. Magnetic resonance imaging was performed for lesions larger than 20 mm. The indications for surgical excision were: hormone activity, lesion diameter more than 30 mm, lesion growth or other radiological features suspicious of malignancy. RESULTS: Of 534 patients assessed for eligibility, 226 (mean age 67 years, 62·4 per cent women; mean lesion diameter 23·9 mm, 22·6 per cent bilateral) fulfilled the inclusion criteria. Mean follow-up was 19·0 months. After baseline evaluation, 14 patients had surgery owing to primary hyperaldosteronism (3), catecholamine-producing tumour (1), tumour size (6), size and indication of subclinical hypercortisolism (3) and metastasis (1). No hypersecreting lesions were confirmed during follow-up; one patient underwent adrenalectomy for a suspected phaeochromocytoma (adrenocortical adenoma at histopathology). No primary adrenal malignancy was found. CONCLUSION: In this prospective cohort study 6·6 per cent of patients with an AI had surgery and benign hormone-producing tumours were verified in 3·1 per cent. Repeat CT and hormone evaluation after 2 years did not increase the sensitivity for diagnosis of malignant or hormone-producing tumours.

PDGF-C is a new protease-activated ligand for the PDGF alpha-receptor.

Platelet-derived growth factors (PDGFs) are important in many types of mesenchymal cell. Here we identify a new PDGF, PDGF-C, which binds to and activates the PDGF alpha-receptor. PDGF-C is activated by proteolysis and induces proliferation of fibroblasts when overexpressed in transgenic mice. In situ hybridization analysis in the murine embryonic kidney shows preferential expression of PDGF-C messenger RNA in the metanephric mesenchyme during epithelial conversion. Analysis of kidneys lacking the PDGF alpha-receptor shows selective loss of mesenchymal cells adjacent to sites of expression of PDGF-C mRNA; this is not found in kidneys from animals lacking PDGF-A or both PDGF-A and PDGF-B, indicating that PDGF-C may have a unique function.

Prognostic role of serum thymidine kinase 1 kinetics during neoadjuvant chemotherapy for early breast cancer.

BACKGROUND: Emerging data support the use of thymidine kinase 1 (TK1) activity as a prognostic marker and for monitoring of response in breast cancer (BC). The long-term prognostic value of TK1 kinetics during neoadjuvant chemotherapy is unclear, which this study aimed to elucidate. METHODS: Material from patients enrolled to the single-arm prospective PROMIX trial of neoadjuvant epirubicin, docetaxel and bevacizumab for early BC was used. Ki67 in baseline biopsies was assessed both centrally and by automated digital imaging analysis. TK1 activity was measured from blood samples obtained at baseline and following two cycles of chemotherapy. The associations of TK1 and its kinetics as well as Ki67 with event-free survival and overall survival (OS) were evaluated using multivariable Cox regression models. RESULTS: Central Ki67 counting had excellent correlation with the results of digital image analysis (r = 0.814), but not with the diagnostic samples (r = 0.234), while it was independently prognostic for worse OS [adjusted hazard ratio (HRadj) = 2.72, 95% confidence interval (CI) 1.19-6.21, P = 0.02]. Greater increase in TK1 activity after two cycles of chemotherapy resulted in improved event-free survival (HRadj = 0.50, 95% CI 0.26-0.97, P = 0.04) and OS (HRadj = 0.46, 95% CI 0.95, P = 0.04). There was significant interaction between the prognostic value of TK1 kinetics and Ki67 (pinteraction 0.04). CONCLUSION: Serial measurement of serum TK1 activity during neoadjuvant chemotherapy provides long-term prognostic information in BC patients. The ease of obtaining serial samples for TK1 assessment motivates further evaluation in larger studies.

Cellular tropism and transduction properties of seven adeno-associated viral vector serotypes in adult retina after intravitreal injection.

Recombinant adeno-associated virus (rAAV) vectors are increasingly being used as tools for gene therapy, and clinical trials have begun in patients with genetically linked retinal disorders. Intravitreal injection is optimal for the transduction of retinal ganglion cells (RGCs), although complete selectivity has not been achieved. There may also be advantages in using intravitreal approaches for the transduction of photoreceptors. Here we compared the cellular tropism and transduction efficiency of rAAV2/1, -2/2, -2/3, -2/4, -2/5, -2/6 and -2/8 in adult rat retina after intravitreal injection. Each vector encoded green fluorescent protein (GFP), and the number, laminar distribution and morphology of transduced GFP(+) cells were determined using fluorescent microscopy. Assessment of transduced cell phenotype was based on cell morphology and immunohistochemistry. rAAV2/2 and rAAV2/6 transduced the greatest number of cells, whereas rAAV2/5 and rAAV2/8 were least efficient. Most vectors primarily transduced RGCs; however, rAAV2/6 had a more diverse tropism profile, with 46% identified as amacrine or bipolar cells, 23% as RGCs and 22% as Müller cells. Müller cells were also frequently transduced by rAAV2/4. The highest photoreceptor transduction was seen after intravitreal rAAV2/3 injection. These data facilitate the design and selection of rAAV vectors to target specific retinal cells, potentially leading to an improved gene therapy for various human retinal pathologies.

Lack of pericytes leads to endothelial hyperplasia and abnormal vascular morphogenesis.

The association of pericytes (PCs) to newly formed blood vessels has been suggested to regulate endothelial cell (EC) proliferation, survival, migration, differentiation, and vascular branching. Here, we addressed these issues using PDGF-B-- and PDGF receptor-beta (PDGFR-beta)--deficient mice as in vivo models of brain angiogenesis in the absence of PCs. Quantitative morphological analysis showed that these mutants have normal microvessel density, length, and number of branch points. However, absence of PCs correlates with endothelial hyperplasia, increased capillary diameter, abnormal EC shape and ultrastructure, changed cellular distribution of certain junctional proteins, and morphological signs of increased transendothelial permeability. Brain endothelial hyperplasia was observed already at embryonic day (E) 11.5 and persisted throughout development. From E 13.5, vascular endothelial growth factor-A (VEGF-A) and other genes responsive to metabolic stress became upregulated, suggesting that the abnormal microvessel architecture has systemic metabolic consequences. VEGF-A upregulation correlated temporally with the occurrence of vascular abnormalities in the placenta and dilation of the heart. Thus, although PC deficiency appears to have direct effects on EC number before E 13.5, the subsequent increased VEGF-A levels may further abrogate microvessel architecture, promote vascular permeability, and contribute to formation of the edematous phenotype observed in late gestation PDGF-B and PDGFR-beta knock out embryos.

Computer-aided detection (CAD) as a second reader using perspective filet view at CT colonography: effect on performance of inexperienced readers.

AIM: To evaluate whether computer-aided detection (CAD) as a second reader using perspective filet view [three-dimensional (3D) filet] improves the performance of inexperienced readers at computed tomography colonography (CTC) compared with unassisted 3D filet and unassisted two-dimensional (2D) CTC. MATERIAL AND METHODS: Fifty symptomatic patients underwent CTC and same-day colonoscopy with segmental unblinding. Two inexperienced readers read the CTC studies on 3D filet and 2D several weeks apart. Four months later, readers re-read the cases only evaluating CAD marks using 3D filet. Suspicious CAD marks not previously described on 3D filet were recorded. Jackknife free-response receiver operating characteristic (JAFROC-1) analysis was used to compare the observers' performances in detecting lesions with 3D filet, 2D and 3D filet with CAD. RESULTS: One hundred and three lesions > or =3mm were detected at colonoscopy with segmental unblinding. CAD alone had a sensitivity of 73% (75/103) at a mean false-positive rate per patient of 12.8 in supine and 11.4 in prone. For inexperienced readers sensitivities with 3D filet with CAD were 58% (60/103) and 48% (50/103) with an improvement of 14-16 percentage points (p<0.05) compared with 2D and of 10-11 percentage points (p<0.05) compared with 3D filet. For inexperienced readers, the false-positive rate was 25-41% and 71-200% higher with 3D filet with CAD compared with 3D filet and 2D, respectively. JAFROC-1 analysis showed no significant differences in per-lesion overall performance among reading modes (p=0.8). CONCLUSION: CAD applied as a second reader using 3D filet increased both sensitivity and the number of false positives by inexperienced readers compared with 3D filet and 2D, thus not improving overall performance, i.e., the ability to distinguish between lesions and non-lesions.

Primary three-dimensional analysis with perspective-filet view versus primary two-dimensional analysis: evaluation of lesion detection by inexperienced readers at computed tomographic colonography in symptomatic patients.

BACKGROUND: "Perspective-filet view" is a novel three-dimensional (3D) viewing technique for computed tomography colonography (CTC). Studies with experienced readers have shown a sensitivity for perspective-filet view similar to that of 2D or 3D endoluminal fly-through in detection of colorectal lesions. It is not known whether perspective-filet view, compared to axial images, improves lesion detection by inexperienced readers. PURPOSE: To compare primary 3D analysis using perspective-filet view (3D Filet) with primary 2D analysis, as used by inexperienced CTC readers. Secondary aims were to compare lesion detection by 3D Filet when used by experienced and inexperienced readers, and to evaluate the effect of combined 3D Filet + 2D analysis. MATERIAL AND METHODS: Fifty symptomatic patients were prospectively enrolled. An experienced reader performed 3D Filet analysis followed by complete 2D analysis (3D Filet + 2D), before colonoscopy with segmental unblinding. Two inexperienced readers (readers 2 and 3), blinded to CTC and colonoscopy findings, retrospectively performed 3D Filet analysis and, after 5 weeks, 2D analysis. True positives >or=6 mm detected by the inexperienced readers with 3D Filet and/or 2D were combined to obtain 3D Filet + 2D. RESULTS: Colonoscopy revealed 116 lesions: 16 lesions >or=10 mm, 19 lesions 6-9 mm, and 81 lesions or=6 mm with 3D Filet and 3D Filet + 2D were 77% and 83%, respectively. For the inexperienced readers, sensitivities for lesions >or=6 mm with 3D Filet and 2D were 51% and 57% (reader 2) and 40% and 43% (reader 3), respectively. There was no significant difference between 3D Filet and 2D regarding sensitivity and reading time. For lesions >or=6 mm, 3D Filet + 2D improved the sensitivity of reader 2 to 63% and of reader 3 to 51%. CONCLUSION: Lesion detection by inexperienced readers using perspective-filet view is comparable to that obtained by 2D. Lesion detection improves by combining 3D Filet + 2D, but not to the level of an experienced reader.

Evaluation of the response to treatment in patients with idiopathic achalasia by the timed barium esophagogram: results from a randomized clinical trial.

To choose which treatment would be most effective for the individual patient with newly diagnosed achalasia is difficult for the tending physician. A diagnostic tool that would allow prediction of the symptomatic and functional response after treatment for achalasia is therefore needed. The timed barium esophagogram (TBE) is a method that allows objective assessment of esophageal emptying, but the value of TBE in the clinical management of achalasia remains to be clarified. The aim of this study was first, to assess the ability of TBE to predict symptoms and treatment failure during post-treatment follow-up. Second, to determine whether esophageal emptying as assessed by TBE differs after treatment with pneumatic dilatation or laparoscopic myotomy. Fifty-one patients with newly diagnosed achalasia were prospectively randomized to pneumatic dilatation (n = 26) or laparoscopic myotomy (n = 25). Evaluation with TBE was performed before (n = 46) and after treatment (n = 43). The median interval between treatment and post-treatment TBE was 6 months, and the median follow-up time after the post-treatment TBE was 18 months. Following therapeutic intervention, TBE parameters did not differ significantly between treatment groups. However, significant correlations were found between the height of the barium column at 1 min and the symptom scores at the end of follow up for 'dysphagia for liquids' (P < 0.05, rho = 0.47), 'chest pain' (P < 0.05, rho = 0.42), and the 'Watson dysphagia score' (P < 0.05, rho = 0.46). Patients with less than 50% improvement in this TBE-parameter (height at 1 min) post-treatment had a 40% risk of treatment failure during follow-up. In summary, pneumatic balloon dilatation and laparoscopic myotomy similarly affected esophageal function as assessed by TBE-emptying. Lack of improvement in barium-column height post-treatment was associated with an increased risk of treatment failure which should motivate close surveillance in order to detect symptomatic recurrence at an early stage.

Additive effects of the herbicide glyphosate and elevated temperature on the branched coral Acropora formosa in Nha Trang, Vietnam.

The combined effects of the herbicide glyphosate and elevated temperature were studied on the tropical staghorn coral Acropora formosa, in Nha Trang bay, Vietnam. The corals were collected from two different reefs, one close to a polluted fish farm and one in a marine-protected area (MPA). In the laboratory, branches of the corals were exposed to the herbicide glyphosate at ambient (28 °C) and at 3 °C elevated water temperatures (31 °C). Effects of herbicide and elevated temperature were studied on coral bleaching using photography and digital image analysis (new colorimetric method developed here based on grayscale), chlorophyll a analysis, and symbiotic dinoflagellate (Symbiodinium, referred to as zooxanthellae) counts. All corals from the MPA started to bleach in the laboratory before they were exposed to the treatments, indicating that they were very sensitive, as opposed to the corals collected from the more polluted site, which were more tolerant and showed no bleaching response to temperature increase or herbicide alone. However, the combined exposure to the stressors resulted in significant loss of color, proportional to loss in chlorophyll a and zooxanthellae. The difference in sensitivity of the corals collected from the polluted site versus the MPA site could be explained by different symbiont types: the resilient type C3u and the stress-sensitive types C21 and C23, respectively. The additive effect of elevated temperatures and herbicides adds further weight to the notion that the bleaching of coral reefs is accelerated in the presence of multiple stressors. These results suggest that the corals in Nha Trang bay have adapted to the ongoing pollution to become more tolerant to anthropogenic stressors, and that multiple stressors hamper this resilience. The loss of color and decrease of chlorophyll a suggest that bleaching is related to concentration of chloro-pigments. The colorimetric method could be further fine-tuned and used as a precise, non-intrusive tool for monitoring coral bleaching in situ.

Optimal Solution Volume for Luminal Preservation: A Preclinical Study in Porcine Intestinal Preservation.

BACKGROUND: Rodent studies suggest that luminal solutions alleviate the mucosal injury and prolong intestinal preservation but concerns exist that excessive volumes of luminal fluid may promote tissue edema. Differences in size, structure, and metabolism between rats and humans require studies in large animals before clinical use. METHODS: Intestinal procurement was performed in 7 pigs. After perfusion with histidine-tryptophan-ketoglutarate (HTK), 40-cm-long segments were cut and filled with 13.5% polyethylene glycol (PEG) 3350 solution as follows: V0 (controls, none), V1 (0.5 mL/cm), V2 (1 mL/cm), V3 (1.5 mL/cm), and V4 (2 mL/cm). Tissue and luminal solutions were sampled after 8, 14, and 24 hours of cold storage (CS). Preservation injury (Chiu score), the apical membrane (ZO-1, brush-border maltase activity), and the electrolyte content in the luminal solution were studied. RESULTS: In control intestines, 8-hour CS in HTK solution resulted in minimal mucosal changes (grade 1) that progressed to significant subepithelial edema (grade 3) by 24 hours. During this time, a gradual loss in ZO-1 was recorded, whereas maltase activity remained unaltered. Moreover, variable degrees of submucosal edema were observed. Luminal introduction of high volumes (2 mL/mL) of PEG solution accelerated the development of the subepithelial edema and submucosal edema, leading to worse histology. However, ZO-1 was preserved better over time than in control intestines (no luminal solution). Maltase activity was reduced in intestines receiving luminal preservation. Luminal sodium content decreased in time and did not differ between groups. CONCLUSIONS: This PEG solution protects the apical membrane and the tight-junction proteins but may favor water absorption and tissue (submucosal) edema, and luminal volumes >2 mL/cm may result in worse intestinal morphology.

Safety, Acceptability and Adherence of Dapivirine Vaginal Ring in a Microbicide Clinical Trial Conducted in Multiple Countries in Sub-Saharan Africa.

BACKGROUND: This was the first microbicide trial conducted in Africa to evaluate an antiretroviral-containing vaginal ring as an HIV prevention technology for women. OBJECTIVES: The trial assessed and compared the safety, acceptability and adherence to product use of a 4-weekly administered vaginal ring containing the antiretroviral microbicide, dapivirine, with a matching placebo ring among women from four countries in sub-Saharan Africa. METHODS: 280 Healthy, sexually active, HIV-negative women, aged 18 to 40 years were enrolled with 140 women randomised to a dapivirine vaginal ring (25 mg) and 140 women to a matching placebo ring, inserted 4-weekly and used over a 12-week period. Safety was evaluated by pelvic examination, colposcopy, clinical laboratory assessments, and adverse events. Blood samples for determination of plasma concentrations of dapivirine were collected at Weeks 0, 4 and 12. Residual dapivirine levels in returned rings from dapivirine ring users were determined post-trial. Participant acceptability and adherence to ring use were assessed by self-reports. RESULTS: No safety concerns or clinically relevant differences were observed between the dapivirine and placebo ring groups. Plasma dapivirine concentrations immediately prior to ring removal were similar after removal of the first and third ring, suggesting consistent ring use over the 12-week period. No clear relationship was observed between the residual amount of dapivirine in used rings and corresponding plasma concentrations. Self-reported adherence to daily use of the vaginal rings over the 12-week trial period was very high. At the end of the trial, 96% of participants reported that the ring was usually comfortable to wear, and 97% reported that they would be willing to use it in the future if proven effective. CONCLUSIONS: The dapivirine vaginal ring has a favourable safety and acceptability profile. If proven safe and effective in large-scale trials, it will be an important component of combination HIV prevention approaches for women. TRIAL REGISTRATION: ClinicalTrials.gov NCT01071174.

Fever, bacteriuria and concomitant disease in children with urinary tract infection.

A total of 124 children aged 0.2 to 6 years were enrolled in a study of first time febrile urinary tract infection. The patient population was stratified in groups according to the stringency of criteria for fever and bacteriuria and the presence of concomitant disease. The major group of 88 patients consisted of children with fever greater than or equal to 38.5 degrees C measured at the hospital within 24 hours of diagnosis, bacteriuria verified by suprapubic bladder aspiration or repeated cultures of voided urine, but without concomitant disease. These children were mainly infected with attaching Escherichia coli specific for galactose alpha (1----4) beta galactose containing receptors and had laboratory evidence of inflammation. Another group of 11 children were distinguished with strictly defined bacteriuria and concomitant disease. These children were infected with nonattaching bacteria and had lower concentrations of C-reactive protein in serum and lower microsedimentation rates than the major group. Five of these children had a reduction in renal concentrating capacity. The study emphasizes the heterogeneity among patients with fever and bacteriuria but does not rule out the possibility of renal involvement in any subgroup.

Pharmacokinetics of peptichemio in myeloma patients: release of m-L-sarcolysin in vivo and in vitro.

Peptichemio (PTC) is a mixture of six synthetic oligopeptides, each of which contains the alkylating residue m-[di(2-chloroethyl)amino]-L-phenylalanine (L-mSL). The fate of PTC was investigated in eight patients with multiple myeloma after intravenous infusion of the drug. The quantitative analysis of the plasma samples was performed by liquid chromatography with fluorometric detection. L-mSL was rapidly released from the peptides and reached its maximal plasma concentration at the end of the infusion. Its median elimination half-life was 1.73 (range, 0.72-2.41) h. It was possible to follow the concentration of only one of the peptides, L-mSL-L-Arg(NO2)-L-Nval.OEt, during and shortly after the infusion of PTC. The stability of L-mSL and the peptides was studied in buffer solution (pH 7.3), plasma, and blood. The stability of some of the peptides was drastically decreased in blood, the degradation half-lives being only about 1 min. We conclude that L-mSL plays an important role in the mechanism of action of PTC.

Comparison of various D-dimer tests for the diagnosis of deep venous thrombosis.

Analyses of D-dimers in plasma are frequently used as diagnostic tools for deep venous thrombosis (DVT). Enzyme-linked immunosorbent assays (ELISAs) are considered to be the method of choice for quantitative assays, but are time consuming. Therefore, we have assessed plasma levels of D-dimers in patients with clinically suspected DVT using quantitative (Asserachrom D-Di ELISA and TintElize), semiquantitative (Minutex latex, D-Di latex, NycoCard D-Dimer) and qualitative (INSTANT.I.A) assays. Phlebography was used as the gold standard to verify or exclude the suspected diagnosis. We conclude that the fast assays, INSTANT.I.A and Minutex, have essentially the same negative predictive value [91% and 89%, respectively, using a cut-off value < 0.5 mg/l fibrinogen equivalent units (FEU)] for excluding DVT as the Asserachrom D-Di ELISA and TintElize tests (92%). The D-Di Latex assay had a negative predictive value of 82% (cut-off < 0.5 mg/l FEU) and turned out to be less useful in our material. The NycoCard D-dimer assay had a negative predictive value of 100% when using the cut-off value < 0.5 mg/l FEU, but this was substantially lower when the cut-off was changed to < or = 0.5 mg/l. Thus, we conclude that several fast tests offer a simpler and more rapid way of determining plasma levels of D-dimer than conventional ELISA methods without loss of clinical usefulness in excluding DVT.