Functionalization of Liposomes with Hydrophilic Polymers Results in Macrophage Uptake Independent of the Protein Corona.

Liposomes are established drug carriers that are employed to transport and deliver hydrophilic drugs in the body. To minimize unspecific cellular uptake, nanocarriers are commonly modified with poly(ethylene glycol) (PEG), which is known to minimize unspecific protein adsorption. However, to date, it has not been studied whether this is an intrinsic and specific property of PEG or if it can be transferred to hyperbranched polyglycerol (hbPG) as well. Additionally, it remains unclear if the reduction of unspecific cell uptake is independent of the "basic" carrier at which a surface functionalization with polymers is usually applied. Therefore, we studied the protein corona of differently functionalized liposomes (unfunctionalized vs PEG or hbPG-functionalized) using PEGylated and PGylated lipids. Their cellular uptake in macrophages was compared. For all three liposomal samples, rather similar protein corona compositions were found, and also-more importantly-the total amount of proteins adsorbed was very low compared to other nanoparticles. Interestingly, the cellular uptake was then significantly changed by the surface functionalization itself, despite the adsorption of a small amount of proteins: although the PEGylation of liposomes resulted in the abovementioned decreased cell uptake, functionalization with hbPG lead to enhanced macrophage interaction-both in the media with and without proteins. In comparison to other nanocarrier systems, this seems to be a liposome-specific effect related to the low amount of adsorbed proteins.

Orthogonal Click Conjugation to the Liposomal Surface Reveals the Stability of the Lipid Anchorage as Crucial for Targeting.

Synthetic access to multiple surface decorations are a bottleneck in the development of liposomes for receptor mediated targeting. This opens a complex multiparameter space, exploration of which is severely limited in terms of sample numbers and turnaround times. Here, we unlock this technological barrier by a combination of a milligram-scale liposome formulation using dual centrifugation and orthogonal click chemistry on the liposomal surface. Application of these techniques to conceptually new amphiphilic compounds, which feature norbornene and alkyne groups at the apex of sterically stabilizing, hyperbranched polyglycerol moieties, revealed a particular influence of the membrane anchor of functional amphiphiles. Folic acid residues clicked to cholesterol-based amphiphiles were inefficient in folate-mediated cell targeting, while dialkyl-anchored amphiphiles remained stable in the liposomal membrane and imparted efficient targeting properties. These findings are of specific importance considering the popularity of cholesterol as a lipophilic anchor.

The importance of glutathione and phytochelatins on the selenite and arsenate detoxification in Arabidopsis thaliana.

We investigated the role of glutathione (GSH) and phytochelatins (PCs) on the detoxification of selenite using Arabidopsis thaliana. The wild-type (WT) of Arabidopsis thaliana and its mutants (glutathione deficient Cad 2-1 and phytochelatins deficient Cad 1-3) were separately exposed to varying concentrations of selenite and arsenate and jointly to both toxicants to determine their sensitivities. The results of the study revealed that, the mutants were about 20-fold more sensitive to arsenate than the WT, an indication that the GSH and PCs affect arsenate detoxification. On the contrary, the WT and both mutants showed a similar level of sensitivity to selenite, an indication that the GSH and PCs do not significantly affect selenite detoxification. However, the WT is about 8 times more sensitive to selenite than to arsenate, and the mutants were more resistant to selenite than arsenate by a factor of 2. This could not be explained by the accumulation of both elements in roots and shoots in exposure experiments. The co-exposure of the WT indicates a synergistic effect with regards to toxicity since selenite did not induce PCs but arsenic and selenium compete in their PC binding as revealed by speciation analysis of the root extracts using HPLC-ICP-MS/ESI-MS. In the absence of PCs an antagonistic effect has been detected which might suggest indirectly that the formation of Se glutathione complex prevent the formation of detrimental selenopeptides. This study, therefore, revealed that PC and GSH have only a subordinate role in the detoxification of selenite.

Classification of the Aging Lips: A Foundation for an Integrated Approach to Perioral Rejuvenation.

BACKGROUND: Although perioral aging is highly individual with several distinct processes taking part simultaneously, there is scarce systematic information which helps to indicate the right rejuvenation approach among the multitude of proposed procedures. Existing data about perioral aging has not yet been transformed into a consistent therapeutic concept. The intention of this study was to provide a simple, yet reproducible classification and to offer appropriate rejuvenation approaches. METHODS: To identify reliable and constant landmarks of the ongoing process of perioral aging, 462 perioral photo documentations were morphometrically analyzed. Based upon the identified landmarks a two-dimensional classification was developed. The classification was validated by three plastic surgeons. Inter- and intra-rater reliability was calculated using Cohen's kappa coefficient. RESULTS: Perioral aging can be broken down into changes of the lip shape and changes of the lip surface. Both processes can be classified into three stages each: Lip shape according to the shape in profile view, the lip length in relation to the frontal incisors, and the degree of vermilion inversion. Lip surface according to the presence and degree of radial wrinkles and the visibility of the structural elements Cupid's bow, philtrum, and white roll. Inter-observer reliability was rated very good (kappa values between 0.819 and 0.963) and perfect for intra-observer reliability (1.0). CONCLUSION: A better understanding of perioral aging leads to a simple classification for the aging lips. Using the classification helps to tailor an appropriate treatment to the individual patient and aids to achieve a natural rejuvenation result.

Neoadjuvant chemotherapy in breast cancer significantly reduces number of yielded lymph nodes by axillary dissection.

BACKGROUND: Neoadjuvant chemotherapy (NC) is an established therapy in breast cancer, able to downstage positive axillary lymph nodes, but might hamper their detectibility. Even if clinical observations suggest lower lymph node yield (LNY) after NC, data are inconclusive and it is unclear whether NC dependent parameters influence detection rates by axillary lymph node dissection (ALND). METHODS: We analyzed retrospectively the LNY in 182 patients with ALND after NC and 351 patients with primary ALND. Impact of surgery or pathological examination and specific histomorphological alterations were evaluated. Outcome analyses regarding recurrence rates, disease free (DFS) and overall survival (OS) were performed. RESULTS: Axillary LNY was significantly lower in the NC in comparison to the primary surgery group (median 13 vs. 16; p < 0.0001). The likelihood of incomplete axillary staging was four times higher in the NC group (14.8% vs. 3.4%, p < 0.0001). Multivariate analyses excluded any influence by surgeon or pathologist. However, the chemotherapy dependent histological feature lymphoid depletion was an independent predictive factor for a lower LNY. Outcome analyses revealed no significant impact of the LNY on local and regional recurrence rates as well as DFS and OS, respectively. CONCLUSION: NC significantly reduces the LNY by ALND and has profound effects on the histomorphological appearance of lymph nodes. The current recommendations for a minimum removal of 10 lymph nodes by ALND are clearly compromised by the clinically already established concept of NC. The LNY of less than 10 by ALND after NC might not be indicative for an insufficient axillary staging.

Reduced Levels of Brain-Derived Neurotrophic Factor Affect Body Weight, Brain Weight and Behavior.

Neurotrophins, which belong to the family of growth factors, not only play crucial roles during development but are also involved in many processes in the postnatal brain. One representative of neurotrophins is brain-derived neurotrophic factor (BDNF). BDNF plays a role in the regulation of body weight and neuronal plasticity and is, therefore, also involved in processes associated with learning and memory formation. Many of the studies on BDNF have been carried out using BDNF-deficient mice. Unfortunately, homozygous deletion of BDNF is lethal in the early postnatal stage, so heterozygous BDNF-deficient mice are often studied. Another possibility is the use of conditional BDNF-deficient mice in which the expression of BDNF is strongly downregulated in some brain cells, for example, in the neurons of the central nervous system, but the expression of BDNF in other cells in the brain is unchanged. To further reduce BDNF expression, we crossed heterozygous BDNF-deficient mice with mice carrying a deletion of BDNF in neurofilament L-positive neurons. These offspring are viable, and the animals with a strong reduction in BDNF in the brain show a strongly increased body weight, which is accompanied by a reduction in brain weight. In addition, these animals show behavioral abnormalities, particularly with regard to locomotion.

Prophylaxis of intra- and postoperative nausea and vomiting in patients during cesarean section in spinal anesthesia.

BACKGROUND: This paper describes a randomized prospective study conducted in 308 patients undergoing caesarean section in spinal anaesthesia at a single hospital between 2010 and 2012 to find a suitable anti-emetic strategy for these patients. MATERIAL AND METHODS: Spinal anesthesia was performed in left prone position, at L3/L4 with hyperbaric 0.5% Bupivacaine according to a cc/cm body height ratio. There were no opioids given peri-operatively. The patients received either no prophylaxis (Group I) or tropisetron and metoclopramide (Group II) or dimenhydrinate and dexamethasone (Group III), or tropisetron as a single medication (Group IV). The primary outcome was nausea and/or vomiting (NV) in the intraoperative, early (0-2 h) or late (2-24 h) postoperative period. Multivariate statistical analysis was conducted with a regression analysis and a backward elimination of factors without significant correlation. RESULTS: All prophylactic agents significantly reduced NV incidence intraoperatively. Relative risk reduction for NV by prophylaxis was most effective (59.5%) in Group II (tropisetron and metoclopramide). In Group III (dimenhydrinate and dexamethasone), NV risk was reduced by 29.9% and by 28.7% in Group IV (tropisetron mono-therapy). The incidence of NV in the early (0?2 h) and the late (2?24 h) postoperative period was low all over (7.8%), but the relative risk reduction of NV in the early postoperative period was 54.1% (Group IV), 45.1% (Group III), and 34.8% (Group II), respectively. In the late postoperative period, there was no significant difference between the 4 groups. CONCLUSIONS: We recommend a prophylactic medication with tropisetron 2 mg and metoclopramide 20 mg for patients during caesarean section. These agents are safe, reasonably priced, and highly efficient in preventing nausea and vomiting.

Mutually distinguishing microRNA signatures of breast, ovarian and endometrial cancers in vitro.

Early diagnosis and therapy in the first stages of a malignant disease is the most crucial factor for successful cancer treatment and recovery. Currently, there is a high demand for novel diagnostic tools that indicate neoplasms in the first or pre­malignant stages. MicroRNAs (miRNA or miR) are small non­coding RNAs that may act as oncogenes and downregulate tumor­suppressor genes. The detection and mutual discrimination of the three common female malignant neoplasia types breast (BC), ovarian (OC) and endometrial cancer (EC) could be enabled by identification of tumor entity­specific miRNA expression differences. In the present study, the relative expression levels of 25 BC, EC and OC­related miRNAs were assessed by reverse transcription­quantitative PCR and determined using the 2­ΔΔCq method for normalization against the mean of four housekeeping genes. Expression levels of all miRNAs were analyzed by regression against cell line as a factor. An expression level­based discrimination between BC and OC cell types was obtained for a subgroup of ten different miRNA types. miR­30 family genes, as well as three other miRNAs, were found to be uniformly upregulated in OC cells compared with BC cells. BC and EC cells could be distinguished by the expression profiles of six specific miRNAs. In addition, four miRNAs were differentially expressed between EC and OC cells. In conclusion, miRNAs were identified as a potential novel tool to detect and mutually discriminate between BC, OC and EC. Based on a subset of 25 clinically relevant human miRNA types, the present study could significantly discriminate between these three female cancer types by means of their expression levels. For further verification and validation of miRNA­based biomarker expression signatures that enable valuable tumor detection and characterization in routine screening or potential therapy monitoring, additional and extended in vitro analyses, followed by translational studies utilizing patients' tissue and liquid biopsy materials, are required.

Morphological, bacterial, and secondary metabolite changes of Aplysina aerophoba upon long-term maintenance under artificial conditions.

The aim of this study was to analyze successional changes in the bacterial community over a period of 6 months of cultivation of Aplysina aerophoba sponges under different artificial cultivation conditions by use of denaturing gradient gel electrophoresis (DGGE). The cultivation conditions varied concerning the water temperature (20 +/- 2 degrees C and 25 +/- 2 degrees C) of the aquaria, additional illumination of one aquarium, and feeding of the sponges. Amplicons from DGGE separation of dominant colonizing or variably appearing bacteria were sequenced and aligned for taxonomical identification. In addition, secondary metabolites typically found in A. aerophoba were analyzed to investigate changes in the natural product profile during cultivation. The cultivation of sponges under any given condition did not lead to a depletion of their bacterial community in the course of the experiment. On the contrary, the distinctive set of associated bacteria was maintained in spite of a dramatic loss of biomass and morphological degradation during the cultivation period. Generally, all sequences obtained from the DGGE gels were related to bacteria of five phyla: Actinobacteria, Cyanobacteria, alpha-Proteobacteria, gamma-Proteobacteria, and Chloroflexi. Despite the overall stability of the bacterial community in A. aerophoba, an unambiguous variability was detected for the Cyanobacteria "A. aerophoba clone TK09". This variability was ascribed to the predominant light conditions. The analysis of the metabolic pattern revealed that the concentration of a class of characteristic-brominated compounds typically found in A. aerophoba, like aeroplysinin-1, aerophobin-1, aerophobin-2, and isofistularin-3, increased over the 6 months of cultivation.

Surface Modification of Nanoparticles and Nanovesicles via Click-Chemistry.

Surface modification of nanocarriers offers the possibility of targeted drug delivery, which is of major interest in modern pharmaceutical science. Click-chemistry affords an easy and fast way to modify the surface with targeting structures under mild reaction conditions. Here we describe our current method for the post-preparational surface modification of multifunctional sterically stabilized (stealth) liposomes via copper-catalyzed azide-alkyne cycloaddition (CuAAC) and inverse electron demand Diels-Alder norbornene-tetrazine cycloaddition (IEDDA). We emphasize the use of these in a one-pot orthogonal reaction for deep investigation on stability and targeting of nanocarriers. As the production of clickable amphiphilic polymers is a limiting factor in most cases, we also describe our nanocarrier preparation technique called dual centrifugation, which enables the formulation of liposomes on a single-digit milligram scale of total lipid mass.

Prophylaxis of postoperative nausea and vomiting in elective breast surgery.

STUDY OBJECTIVE: To evaluate strategies to treat postoperative nausea and vomiting (PONV) in patients undergoing elective breast surgery. DESIGN: Prospective, randomized, double-blinded, placebo-controlled trial. SETTING: University-affiliated hospital. PATIENTS: 480 patients with risk factors for PONV. INTERVENTIONS: Patients were randomized to three groups to receive an antiemetic prophylactic combination of haloperidol and tropisetron (Group HT), dimenhydrinate and dexamethasone (Group DD), or no prophylaxis (Group P). Anesthesia was maintained with volatile anesthesia (desflurane or sevoflurane) and fentanyl or total intravenous anesthesia (TIVA). MEASUREMENTS: Incidence of nausea, emesis, or both in the early (0 - 2 hrs) and late (2 - 24 hrs) postoperative periods were recorded, as were the number of episodes and the time of each occurrence; and patient assessment of the PONV experience on a scale comparable to a numeric rating scale (NRS). MAIN RESULTS: Both antiemetic combinations significantly reduced PONV incidence. In patients who received no prophylaxis, PONV incidence was 48.2% in patients given volatile anesthetics and 43.8% in those who received TIVA. PONV incidence was 17.5% in the Group HT patients who received volatile anesthetics, and 25% in the Group HT patients who received TIVA. PONV incidence was 11.4% in Group DD patients given volatile anesthetics, and 15% in Group DD patients receiving TIVA. TIVA reduced the incidence of PONV in the early postoperative period (0-2 hrs), but increased PONV incidence in the late period (2-24 hrs). Patients given TIVA with propofol and remifentanil intraoperatively required more opioids postoperatively than patients given volatile anesthetics. CONCLUSION: The frequency of PONV was reduced significantly with both antiemetic combinations.

Improved safety of autologous breast reconstruction surgery by stabilisation of microsurgical vessel anastomoses using fibrin sealant in 349 free DIEP or fascia-muscle-sparing (fms)-TRAM flaps: a two-centre study.

BACKGROUND: We routinely perform free DIEP flap and fascia-muscle-sparing (fms) TRAM flap procedures using fibrin sealant to stabilise anastomosed vessels, thus avoiding some of the difficulties associated with microsurgical anastomoses. METHODS: Women undergoing elective, autologous breast reconstruction with free DIEP flaps or fms-TRAM flaps between June 2004 and June 2007 in two Interdisciplinary Breast Centres were included in a retrospective chart review. RESULTS: A total of 349 breast reconstructions were performed in 325 women. Of these, 201 (57.6%) were free DIEP flap procedures and 148 (42.4%) were fms-TRAM flap procedures. Average hospital stay was 9.8 days. Complete flap loss was seen in 3 cases (0.9%). Low rates of post-operative complications were observed. CONCLUSIONS: Microsurgical breast reconstruction using free DIEP flaps and fms-TRAM flaps, with fibrin sealant for stabilisation of microvascular anastomoses, provides good post-operative outcome featuring a low incidence of flap loss or other common post-operative complications.

Breast shaping by an isolated tissue flap.

Breast reshaping surgery for tuberous breast, breast reduction, and mastopexy procedures aim to keep the gained shape for a long time. In breast reduction, the surgeon must avoid loss of fullness in the upper pole, descent of the breast mass known as secondary dropout, and relapsed shape of the repaired tuberous breasts. As described in their clinical report, the authors use a caudally based thoracic wall flap to avoid these problems. The blood supply to the breast comes from two main sources: the mammary internal and lateral arteries. Because of vessels constantly perforating the pectoralis major muscle, it is possible to isolate a caudally based thoracic wall flap. These vessels originate from intercostal arteries as anteromedial intercostal perforators, and from the thoracoacromial artery as in the skin paddle of the pectoralis major muscle flap. This flap is long enough to reach every part of the breast where it is needed. Between January 2002 and June 2005, 64 patients underwent procedures in which the caudally based thoracic wall flap was used.