Physico-chemical analysis of electrospun fibers - A systematic approach.

Electrospun fibers emerged as promising drug delivery systems for various pharmaceutical applications due to their favorable properties. However, while for established drug delivery systems (e.g. tablets or capsules) standardized analytical procedures exist, the methodologies for characterization of electrospun fibers differ widely in the literature. Unfortunately, this situation impedes comparison of different studies and consequently hampers translation of the results into clinics. Thus, there is an urgent need for systematic studies evaluating different analytical techniques for their validity to characterize and differentiate different electrospun fibers. In this study, we aimed to identify a predictive and robust toolset of complementary analytical techniques allowing for comprehensive and discriminative evaluation of electrospun fibers. For this purpose, we fabricated two drug-loaded model formulations with contrastive physico-chemical properties and drug release kinetics. Different analytical techniques were applied for physico-chemical characterization of the spinning solutions as well as of the fibers. Each analytical method was evaluated with regard to discriminative power and individual limitations. The introduction of novel analytical approaches such as automated low-volume release testing may further advance the field of electrospinning. By combining complementary analytical methods, including spectral composition analysis, morphology visualization, characterization of physico-chemical properties and drug release kinetics, as well as the application of multivariate data analysis, we were able to establish a robust and predictive toolset, which can support comparability of future electrospinning studies and the translation from the lab bench into clinics.

In-situ tear fluid dissolving nanofibers enable prolonged viscosity-enhanced dual drug delivery to the eye.

Liquid and semi-solid formulations are the most commonly used drug delivery systems for ophthalmic diseases. Upon application into the conjunctival sac, these systems introduce a variable and unphysiologically high liquid volume to the eye, resulting in overflow and extensive nasolacrimal drainage, accounting for dosing inaccuracy and short ocular residence time. In this study, we present nanofibrous electrospun scaffolds composed of biocompatible polymers, overcoming these challenges by immediate drug release. The fibers incorporate gentamicin and dexamethasone, intended for the treatment of bacterial conjunctivitis. Upon contact with the ocular surface, the nanofibers immediately dissolve in the tear fluid, quantitatively releasing the two actives, yielding over92% drug recovery, determined with fluorimetric and chromatographic quantifications methods. Simultaneously, the viscosity of the tear fluid increases, shown by complex viscometry measurements. A newly developed ex vivo microfluidic porcine cornea model was used to evaluated ocular residence time. In contrast to fluid eye drops, the contact time was significantly prolonged and 20 min after application, an increase in drug availability on the ocular surface of 342% was observed. Biocompatibility of the polymer system was demonstrated in an OECD approved in vitro cornea model. The antibacterial activity after processing was evaluated according to EUCAST guidelines, and storage stability of the system was confirmed over a 12-week period. This innovative drug delivery system poses a highly promising platform technology, overcoming challenges associated with conventional dosage forms for drug delivery to the anterior eye and thus significantly advancing therapeutic approaches.

The functional relevance of passenger leukocytes and microchimerism for heart allograft acceptance in the rat.

With an organ transplant, hematopoietic donor cells are transferred to the recipient. To study the relevance of the resulting microchimerism for allograft acceptance, we analyzed a rat model of cyclosporine-induced tolerance for strongly incompatible heart allografts. Using a monoclonal antibody that detects a donor-specific CD45 allotype (RT7a), we selectively depleted donor leukocytes at different times after transplantation (days 0 or 18). Depletion was similarly effective at both times. However, only depletion on day 0 prevented tolerance induction and was associated with severe acute or chronic graft rejection. This indicates that passenger leukocytes have an essential immunomodulatory effect on the induction phase of allograft acceptance.

Bandwidth-tunable single-photon source in an ion-trap quantum network.

We report a tunable single-photon source based on a single trapped ion. Employing spontaneous Raman scattering and in-vacuum optics with large numerical aperture, single photons are efficiently created with controlled temporal shape and coherence time. These can be varied between 70 ns and 1.6 micros, as characterized by operating two sources simultaneously in two remote ion traps which reveals mutual and individual coherence through two-photon interference.

Improvement in long-term renal graft survival due to CMV prophylaxis with oral ganciclovir: results of a randomized clinical trial.

Oral ganciclovir prophylaxis and intravenous preemptive therapy are competitive approaches to prevent cytomegalovirus (CMV) disease after renal transplantation. This trial compared efficacy, safety and long-term graft outcome in 148 renal graft recipients randomized to ganciclovir prophylaxis (N = 74) or preemptive therapy (N = 74). Hierarchical testing revealed (i) patients with CMV infection had more severe periods of impaired graft function (creatinine clearance(max-min) 25.0 +/- 14.2 mL/min vs. 18.1 +/- 12.5 mL/min for patients without CMV infection; p = 0.02),(ii) prophylaxis reduced CMV infection by 65% (13 vs. 33 patients; p < 0.0001) but (iii) creatinine clearance at 12 months was comparable for both regimes (54.0 +/- 24.9 vs. 53.1 +/- 23.7 mL/min; p = 0.92). No major safety issues were observed, and patient survival at 12 months was similar in both groups (5 deaths [6.8%] vs. 4 [5.4%], p = 1.0000). Prophylaxis significantly increased long-term graft survival 4 years posttransplant (92.2% vs. 78.3%; p = 0.0425) with a number needed to treat of 7.19. Patients with donor +/recipient + CMV serostatus had the lowest rate of graft loss following prophylaxis (0.0% vs. 26.8%; p = 0.0035). In conclusion, it appears that routine oral prophylaxis may improve long-term graft survival for most renal transplant patients. Preemptive therapy can be considered in low risk patients in combination with adequate CMV monitoring.

Long-term allograft acceptance induced by single dose anti-leukocyte common antigen (RT7) antibody in the rat.

BACKGROUND: In clinical organ transplantation monoclonal antibodies (mAb) to different surface molecules of immunocompetent cells become integral parts of the immunosuppressive therapy. In this study, a mAb against the rat leukocyte common antigen CD45 (RT7) was tested for its immunosuppressive potency after a single perioperative injection. METHODS: Binding and depleting properties of the anti-RT7 mAb were investigated by flow cytometry. In the fully major histocompatibility complex-disparate heart and skin transplantation models (LEW [RT1l]--> LEW.1W [RT1u]), a single dose of anti-RT7 mAb (10 mg/kg) was administered intravenously (day -1). To characterize the long-term acceptance of heart allografts second set skin transplantation (day 100), mixed lymphocyte reaction studies (day 100) and reverse transcriptase-polymerase chain reaction analysis for intragraft cytokine expression (day 200) were performed. RESULTS: The anti-RT7 mAb bound to nearly all hematopoietic lineage cells, but particularly T and NK cells, and profoundly depleted these cells in circulation and lymphoid tissues. Anti-RT7 mAb-treated rats showed long-term acceptance of heart allografts (>200 days; n=12), whereas untreated recipients rejected allografts by day 8 (n=6). In contrast to hearts, primary skin allograft survival was only moderately prolonged. Animals with stable heart allograft acceptance showed normal in vitro lymphocyte proliferation responses to donor and third party antigen. These recipients also acutely rejected second set donor-strain skin grafts without inducing rejection of persisting heart allografts. Reverse transcriptase-polymerase chain reaction analysis of intragraft cytokines showed up-regulation of Fas-ligand and IL-4 mRNA in long-surviving heart allografts. CONCLUSIONS: The findings demonstrate that a single injection of an anti-RT7 mAb in the rat can induce stable long-term acceptance of heart allografts by transient but profound T-cell depletion. Local immunoregulatory mechanisms seem to play a role for maintenance of long-term graft acceptance.

Glycine reduces the inflammatory response and organ damage in a two-hit sepsis model in rats.

The goal of this study was to investigate whether prefeeding of glycine reduces the immunoinflammatory response, the degree of distant organ injury (liver), and/or the mortality rate in a two-hit model using intestinal ischemia/reperfusion and endotoxin (ET) challenge 6 h later in rats. The liver damage was greatest at 24 h after ET challenge and completely inhibited by glycine. The early systemic increase of the proinflammatory cytokines tumor necrosis factor-alpha (TNF-alpha) and interleukin (IL) -6 as well as the secretion of the antiinflammatory cytokine IL-10 was reduced by glycine. Tissue cytokine mRNA expression (TNF-alpha, IL-1beta, IL-10) was decreased in the lung and the liver but not in the mesenteric lymph node or ileum, in the glycine-fed group. However, glycine did not decrease the mortality rate. These results suggest that prefeeding of glycine reduces liver damage as well as the systemic and local (lung and liver) inflammatory response after intestinal ischemia/reperfusion and endotoxin challenge in rats.

Insula and sensory insular cortex and somatosensory control in patients with insular stroke.

BACKGROUND: In functional imaging studies, the insular cortex (IC) has been identified as an essential part of the processing of a whole spectrum of multimodal sensory input. However, there are no lesion studies including a sufficient number of patients, which would reinforce the functional imaging data obtained from healthy subjects. Such lesion studies should examine how damage to the IC affects sensory perception. We chose acute stroke patients with lesions affecting the IC in order to fill this gap. METHODS: A comprehensive sensory profiling by applying a quantitative sensory testing protocol was performed and a voxel-lesion behaviour mapping analysis in 24 patients with acute unilateral cortical damage was applied. RESULTS: Our data demonstrate that patients with lesions of the posterior IC have deficits in temperature perception, but did not show other sensory deficits such as hot or cold pain perception associated with specific lesion locations. CONCLUSION: Our data allow the conclusion that the posterior IC may represent the major region responsible for encoding warm and cold perception in the brain. To what extent focal IC lesions may also impair pain processing or induce post-stroke pain has to be addressed in future studies including more patients.

Thoracic complications of amoebic liver abscess.

We report the cases of 170 patients with thoracic complications from amoebic hepatic abscess. The clinical, radiological and therapeutic features of the condition are discussed. Mortality is high, especially where poverty has already impaired health.

Ketotifen in HIV-infected patients: effects on body weight and release of TNF-alpha.

OBJECTIVE: Tumor necrosis factor alpha (TNF-alpha) is involved in the genesis of HIV-associated malnutrition. We performed an open-label trial on the effects of ketotifen, an in vitro inhibitor of TNF-alpha release from peripheral blood mononuclear cells (PBMCs), on the nutritional status and TNF-alpha release of HIV-infected subjects. PATIENTS: Six HIV-infected subjects received oral ketotifen 4 mg per day for 84 days and were followed up for an additional 70-day period. Body composition was measured by bioelectrical impedance analysis. TNF-alpha plasma levels, TNF-alpha release from PBMCs, and plasma concentration of soluble TNF receptors were measured repeatedly during the study and control period. RESULTS: During ketotifen intake, TNF-alpha release from stimulated PBMCs significantly decreased (68 vs 155 pg ml-1), but not TNF-alpha and soluble TNF receptor plasma concentrations. Subjects gained weight (+ 2.7 kg), whereas weight loss was observed after cessation of treatment (-1.6 kg). CONCLUSION: Ketotifen inhibits TNF-alpha release from stimulated PBMCs and might thus be useful in the management of HIV-associated malnutrition.

Discordant patterns of morphological and molecular change in broadtail madtoms (genus Noturus).

Two morphologically distinct forms of an undescribed madtom catfish (Noturus sp.) occur in the rivers and lakes of southeastern USA. 'Lake' broadtail madtoms are endemic to Lake Waccamaw and are probably related to nearby 'river' broadtail populations. To investigate phylogenetic relationships, we surveyed mitochondrial DNA (mtDNA) sequence variation in 'lake' and 'river' broadtails and other members of the genus Noturus. Mitochondrial rDNA data suggest a sister group relationship between broadtail madtoms and N. insignis, not N. leptacanthus as posited previously. Population-level analyses using additional mtDNA characters (NADH dehydrogenase subunit 2 (ND2) and cytochrome b (Cytb)) identified two highly divergent genetic lineages within broadtail madtoms that do not correspond to the morphological designations 'river' and 'lake'.

Gene-gene concordance and the phylogenetic relationships among rare and widespread pygmy sunfishes (genus Elassoma).

Pygmy sunfishes (Elassoma) are primarily lowland species with an interesting biogeographic dichotomy: three species have broad geographic distributions, and three are narrowly distributed (and have been recommended for threatened or endangered status). To test phylogenetic predictions derived from the geographic distributions of pygmy sunfishes and possible historical factors contributing to the threatened/endangered status of the rare species, we reconstructed trees for two mitochondrial genes and introns of three nuclear genes. The pattern and rate of nuclear and mitochondrial sequence evolution were heterogeneous within Elassoma, but relationships were generally concordant across gene trees. Elassoma is monophyletic and, as predicted by geographic distributions, E. evergladei, E. okefenokee, and E. zonatum consistently branch from deeper nodes. Phylogeographic structure in mitochondrial and nuclear genes also supports an early origin of E. zonatum. Phylogenetic analyses of the five loci support widely divergent positions for the rare species E. alabamae. Two rare species, E. boehlkei and E. okatie, are sister taxa and are related to a widespread species, E. evergladei.

Quantification of alpha 1-fetoprotein mRNA in peripheral blood and bone marrow: a tool for perioperative evaluation of patients with hepatocellular carcinoma.

BACKGROUND/AIMS: Quantification of alpha 1-fetoprotein (AFP) mRNA in the blood using reverse transcriptase polymerase chain reaction (RT-PCR) could be a useful tool in monitoring the dynamics of minimal residual disease in patients with hepatocellular carcinoma (HCC). Since all available assays do not take into account the efficiency of cell separation, RNA extraction and reverse transcription, a competitive RT-PCR assay for quantification of AFP mRNA in relation to the housekeeping gene glyceraldehyde phosphate dehydrogenase (GAPDH) was established. PATIENTS AND METHODS: Peripheral blood of 22 patients and bone marrow aspirates of 11 patients with hepatocellular carcinoma was monitored perioperatively. Eighteen patients with other hepatic tumours or non-malignant hepatic diseases and 26 healthy blood donors served as controls. Messenger RNA contents were calculated relative to the content of GAPDH mRNA as an indicator of total cell count. RESULTS: Among HCC patients, 6 of 22 (26%) were positive for AFP mRNA before operation with values ranging from 2 ag/100 fg to 36 ag/100 fg GAPDH mRNA (mean 14). Among bone marrow samples, AFP mRNA was detectable in 5 of 11 (45%) cases, with 4 ag/100 fg to 23 ag/100 fg GAPDH (mean 9). However, AFP mRNA was also detectable in 3 of 18 (17%) control patients and in 2 of 26 (8%) healthy blood donors. Perioperative findings were highly variable. CONCLUSION: AFP mRNA is not a specific marker for circulating malignant hepatocytes. Whether definition of a cut-off level or the use of a multimarker-PCR will provide more useful data remains to be established.

Body mass index and the prevalence of hypertension and dyslipidemia.

OBJECTIVE: To describe and evaluate relationships between body mass index (BMI) and blood pressure, cholesterol, high-density lipoprotein-cholesterol (HDL-C), and hypertension and dyslipidemia. RESEARCH METHODS AND PROCEDURES: A national survey of adults in the United States that included measurement of height, weight, blood pressure, and lipids (National Health and Nutrition Examination Survey III 1988-1994). Crude age-adjusted, age-specific means and proportions, and multivariate odds ratios that quantify the association between hypertension or dyslipidemia and BMI, controlling for race/ethnicity, education, and smoking habits are presented. RESULTS: More than one-half of the adult population is overweight (BMI of 25 to 29.9) or obese (BMI of > or =30). The prevalence of high blood pressure and mean levels of systolic and diastolic blood pressure increased as BMI increased at ages younger than 60 years. The prevalence of high blood cholesterol and mean levels of cholesterol were higher at BMI levels over 25 rather than below 25 but did not increase consistently with increasing BMI above 25. Rates of low HDL-C increased and mean levels of HDL-C decreased as levels of BMI increased. The associations of BMI with high blood pressure and abnormal lipids were statistically significant after controlling for age, race or ethnicity, education, and smoking; odds ratios were highest at ages 20 to 39 but most trends were apparent at older ages. Within BMI categories, hypertension was more prevalent and HDL-C levels were higher in black than white or Mexican American men and women. DISCUSSION: These data quantify the strong associations of BMI with hypertension and abnormal lipids. They are consistent with the national emphasis on prevention and control of overweight and obesity and indicate that blood pressure and cholesterol measurement and control are especially important for overweight and obese people.