Are charged tips driving TERS-resolution? A full quantum chemical approach.

Experimental evidence suggests an extremely high, possibly even sub-molecular, spatial resolution of tip-enhanced Raman spectroscopy (TERS). While the underlying mechanism is currently still under discussion, two main contributions are considered: The involved plasmonic particles are able to highly confine light to small spatial regions in the near-field, i.e., the electromagnetic effect and the chemical effect due to altered molecular properties of the sample in close proximity to the plasmonic tip. Significant theoretical effort is put into the modeling of the electromagnetic contribution by various groups. In contrast, we previously introduced a computational protocol that allows for the investigation of the local chemical effect-including non-resonant, resonant, and charge transfer contributions-on a plasmonic hybrid system by mapping the sample molecule with a metallic tip model at the (time-dependent) density functional level of theory. In the present contribution, we evaluate the impact of static charges localized on the tip's frontmost atom, possibly induced by the tip geometry in the vicinity of the apex, on the TERS signal and the lateral resolution. To this aim, an immobilized molecule, i.e., tin(II) phthalocyanine (SnPc), is mapped by the plasmonic tip modeled by a single positively vs negatively charged silver atom. The performed quantum chemical simulations reveal a pronounced enhancement of the Raman intensity under non-resonant and resonant conditions with respect to the uncharged reference system, while the contribution of charge transfer phenomena and of locally excited states of SnPc is highly dependent on the tip's charge.

Dynamic inversion enables external magnets to concentrate ferromagnetic rods to a central target.

The ability to use magnets external to the body to focus therapy to deep tissue targets has remained an elusive goal in magnetic drug targeting. Researchers have hitherto been able to manipulate magnetic nanotherapeutics in vivo with nearby magnets but have remained unable to focus these therapies to targets deep within the body using magnets external to the body. One of the factors that has made focusing of therapy to central targets between magnets challenging is Samuel Earnshaw's theorem as applied to Maxwell's equations. These mathematical formulations imply that external static magnets cannot create a stable potential energy well between them. We posited that fast magnetic pulses could act on ferromagnetic rods before they could realign with the magnetic field. Mathematically, this is equivalent to reversing the sign of the potential energy term in Earnshaw's theorem, thus enabling a quasi-static stable trap between magnets. With in vitro experiments, we demonstrated that quick, shaped magnetic pulses can be successfully used to create inward pointing magnetic forces that, on average, enable external magnets to concentrate ferromagnetic rods to a central location.

Tuning of photocatalytic activity by creating a tridentate coordination sphere for palladium.

The synthesis and characterisation of an asymmetric potential bridging ligand bmptpphz (bmptpphz = 2,17-bis(4-methoxyphenyl)tetrapyrido[3,2-a:2',3'-c:3'',2''-h:2''',3'''-j] phenazine) is presented. This ligand contains a 1,10-phenanthroline (phen) and a 2,9-disubstituted phen sphere and possesses a strong absorbance in the visible. Facile coordination of the phen sphere to a Ru(tbbpy)2 core leads to Ru(bmptpphz) ([(tbbpy)2Ru(bmptpphz)](PF6)2; tbbpy = 4,4'-di-tert-butyl-2,2'-bipyridine). UV-vis, emission, resonance Raman and theoretical investigations show that this complex possesses all properties associated with a Ru(tpphz) ([(tbbpy)2Ru(tpphz)](PF6)2; tpphz = tetrapyrido[3,2-a:2',3'-c:3'',2''-h:2''',3'''-j] phenazine) moiety and that the ligand based absorbances in the vis-part also populate an MLCT like state. The coordination of a Pd-core in the new 2,9-disubstituted phen sphere is possible, leading to a cyclometallation. The tridentate complexation leads to changes in the UV-vis and emission behaviour. Furthermore, the stability of the Pd-coordination is significantly enhanced if compared to the unsubstituted Ru(tpphz). Ru(bmptpphz)PdCl proved to be an active photocatalyst for H2 evolution, albeit with lower activity than the mother compound Ru(tpphz)PdCl2.

Antihypertensive efficacy of the angiotensin receptor blocker azilsartan medoxomil compared with the angiotensin-converting enzyme inhibitor ramipril.

Drug therapy often fails to control hypertension. Azilsartan medoxomil (AZL-M) is a newly developed angiotensin II receptor blocker with high efficacy and good tolerability. This double-blind, controlled, randomised trial compared its antihypertensive efficacy and safety vs the angiotensin-converting enzyme inhibitor ramipril (RAM) in patients with clinic systolic blood pressure (SBP) 150-180 mm Hg. Patients were randomised (n=884) to 20 mg AZL-M or 2.5 mg RAM once daily for 2 weeks, then force-titrated to 40 or 80 mg AZL-M or 10 mg RAM for 22 weeks. The primary endpoint was change in trough, seated, clinic SBP. Mean patient age was 57±11 years, 52.4% were male, 99.5% were Caucasian. Mean baseline BP was 161.1±7.9/94.9±9.0 mm Hg. Clinic SBP decreased by 20.6±0.95 and 21.2±0.95 mm Hg with AZL-M 40 and 80 mg vs12.2±0.95 mm Hg with RAM (P<0.001 for both AZL-M doses). Adverse events leading to discontinuation were less frequent with AZL-M 40 and 80 mg (2.4% and 3.1%, respectively) than with RAM (4.8%). These data demonstrated that treatment of stage 1-2 hypertension with AZL-M was more effective than RAM and better tolerated.

Co-adjuvant effects of retinoic acid and IL-15 induce inflammatory immunity to dietary antigens.

Under physiological conditions the gut-associated lymphoid tissues not only prevent the induction of a local inflammatory immune response, but also induce systemic tolerance to fed antigens. A notable exception is coeliac disease, where genetically susceptible individuals expressing human leukocyte antigen (HLA) HLA-DQ2 or HLA-DQ8 molecules develop inflammatory T-cell and antibody responses against dietary gluten, a protein present in wheat. The mechanisms underlying this dysregulated mucosal immune response to a soluble antigen have not been identified. Retinoic acid, a metabolite of vitamin A, has been shown to have a critical role in the induction of intestinal regulatory responses. Here we find in mice that in conjunction with IL-15, a cytokine greatly upregulated in the gut of coeliac disease patients, retinoic acid rapidly activates dendritic cells to induce JNK (also known as MAPK8) phosphorylation and release the proinflammatory cytokines IL-12p70 and IL-23. As a result, in a stressed intestinal environment, retinoic acid acted as an adjuvant that promoted rather than prevented inflammatory cellular and humoral responses to fed antigen. Altogether, these findings reveal an unexpected role for retinoic acid and IL-15 in the abrogation of tolerance to dietary antigens.

Improved culture methods to expand Schwann cells with altered growth behaviour from CMT1A patients.

A duplication of the gene for myelin protein PMP22 is by far the most common cause of the hereditary demyelinating neuropathy CMT1A. A role for PMP22 in cell growth in addition to its function as a myelin protein has been suggested because PMP22 is homologous to a gene specifically upregulated during growth arrest. Furthermore, transfected rat Schwann cells overexpressing PMP22 show reduced growth. In addition, abnormal Schwann cell differentiation has been described in nerve biopsies from CMT1A patients. To analyse whether the duplication of the PMP22 gene in CMT1A neuropathy primarily alters Schwann cell differentiation and to exclude nonspecific secondary responses, we improved human Schwann cell culturing. This allowed us long-term passaging of human Schwann cells with unchanged phenotype, assessed by expression of different Schwann cell markers. Subsequently we established Schwann cell cultures from CMT1A nerve biopsies. We find decreased proliferation of Schwann cells from different CMT1A patients in all passages. We also demonstrate PMP22 mRNA overexpression in cultured CMT1A Schwann cells. We conclude that decreased proliferation in cultured Schwann cells that carry the CMT1A duplication indicates abnormal differentiation of CMT1A Schwann cells. The identification of an abnormal phenotype of CMT1A Schwann cells in culture could possibly lead to an in vitro disease model.

CD44 antibodies inhibit osteoclast formation.

Osteoclast differentiation is a complex process requiring multiple factors and sequential regulation. We have determined that CD44, a cell surface glycoprotein that is known to function as an adhesion receptor, is involved in this process. By immunocytochemistry, we show that CD44 is expressed in mouse osteoclasts that develop in primary cultures of bone marrow cells treated with 1 alpha, 25-dihydroxyvitamin D3. Monoclonal antibodies to CD44 inhibit osteoclast formation in bone marrow cultures in a dose- and time-dependent manner. In contrast, CD44 Fab monomer antibodies have no effect on osteoclast development, suggesting that the inhibition of differentiation by the whole antibodies is facilitated by cross-linking of CD44 molecules. Cocultures of spleen cells and ST2 bone marrow stromal cells indicate that hematopoietic cells mediate the CD44 antibody inhibitory effect. CD44 antibodies do not inhibit osteoclast resorption of calcified matrix, indicating that CD44 is not absolutely required for resorption activity. These observations demonstrate that CD44 may play a role in osteoclast formation and suggest mechanisms by which CD44 antibody effects are mediated.

Genomic sequence of the DAX1 gene: an orphan nuclear receptor responsible for X-linked adrenal hypoplasia congenita and hypogonadotropic hypogonadism.

The gene responsible for X-linked adrenal hypoplasia congenita, DAX1, encodes a member of the nuclear hormone receptor superfamily. We sequenced 8851 bp that contained the DAX1 genomic region. The DAX gene was composed of two exons and one 3.4-kilobase intron. Putative TATA and GC boxes and a putative steroidogenic factor 1 response element were present in the 5'-flanking region. Two potentially polymorphic short tandem repeats were identified. The first exon encoded two putative novel zinc finger motifs within a putative DNA binding domain and part of the ligand binding domain, and the second exon encoded the remainder of the ligand binding domain. Although the putative DNA binding domain of DAX1 does not contain substantial sequence similarity to other nuclear hormone receptor superfamily members, the putative ligand binding domain had remarkable similarity to other family members. Single-strand conformational polymorphism analysis permitted identification of three new mutations in DAX1. In conclusion, single-strand conformational polymorphism analysis facilitates identification of mutations in the DAX1 gene, and the short tandem repeats may permit linkage analysis in families in which mutations are not yet identified. We speculate that DAX1 may be the most primitive member of the nuclear hormone receptor superfamily identified in mammals.

Renal biopsy: why and when.

The purpose of this paper is to emphasize the importance of information obtained by renal biopsy in the diagnosis, prognosis, and therapy of patients with renal disease. Because controversy persists regarding the value of renal biopsy as an aid in determining prognosis and in choosing appropriate therapy, there has been some reluctance to use it early after the onset of obvious signs, symptoms, and laboratory findings indicative of renal disease with or without involvement of other organs. Although all such patients may not benefit from the information provided by a proper biopsy, we will illustrate some of the characteristic histologic details found in specific circumstances in our experience where the biopsy has been particularly helpful in reaching a diagnosis, in assessing prognosis, and in choosing therapy.

Full-lenth cloning, expression and cellular localization of rat plasmolipin mRNA, a proteolipid of PNS and CNS.

We have isolated a 1.476 bp cDNA (NTII11) representing a transcript that is differntially expressed during sciatic nerve development and regeneration in the rat. Nucleotide sequence comparison indicates partial identity with a recently isolated plasmolipin cDNA. However, our clone extends the published sequence by 234 bp at the 5' end and predicts a protein that contains an additional 25 amino acids at th N-terminus. The open reading frame of th NTII11 transcript encodes a 19.4 kDa protein with four putative transmembrane domains. Northern blot analyses revealed a tissue-specific expression was confirmed by in situ hybridization, and cellular localization of plasmolipin mRNA was demonstrated in Schwann cells of the sciatic nerve and in glial cells of myelinated brain structures. The steady-state levels of plasmolipin mRNA were markedly altered (i) during development of sciatic nerve and brain. (ii) after sciatic nerve injury, and (ii) in cured Schwann cells maintained under different conditions of cell growth and arrest. Our data indicate a function of plasmolipin during myelination in the central as well as in the peripheral nervous system.

Prevention of dry socket with clindamycin. A retrospective study.

Clindamycin and other agents were compared for efficacy in preventing the entity "dry socket." A total 765 patients were treated with clindamycin, per os, and 408 patients were treated with other antibiotics or were non-treated controls. All patients underwent surgical removal of impacted mandibular third molars. The incidence of dry socket in untreated control and in non-clindamycin antibiotic-treated patients varied from 15 to 31 percent, while in those patients receiving clindamycin, the incidence was 0.65 percent. The results demonstrate a remarkable effectiveness of clindamycin in reducing the incidence of dry socket following surgical removal of impacted mandibular third molar.

Specificity of simple hormone response elements in androgen regulated genes.

Androgen (AR) and glucocorticoid (GR) receptors recognize a family of 15 base pair partial palindromic hormone response elements (HRE). We have studied receptor interactions with several HREs from androgen regulated genes to determine their potential to mediate a selective androgen response. Synthetic oligonucleotides corresponding to the elements were analysed for receptor binding and steroid dependent transcriptional enhancer activities. Each HRE contained the 3' half-site sequence (5'-TGTNCT-3') of the glucocorticoid response element (GRE) consensus sequence. HREs that countained the 5' half-site GRE consensus sequence (5'-A/GGNACA/G-3') had the strongest and-rogen response element (ARE) and GRE activities. In methylation interference assays, AR and GR interacted with identical base contact sites in the response elements. Two elements that deviated from the GRE consensus sequence by a single optimal base in the 5' half, had reduced ARE activity with no significant change in GRE activity and displayed lower binding of AR than GR in mobility shift assays using purified DNA binding domain peptides. Transfections with AR/GR and GR/AR chimeras containing the N-terminal domain of one receptor linked to the DNA-binding and C-terminal domains of the other suggested that N-terminal domain functions of GR also contributed to the greater GRE than ARE activities of the response elements.

Androgen and glucocorticoid receptors interact with insulin degrading enzyme.

We recently identified a protein, designated receptor accessory factor (RAF), that interacts directly with and enhances the DNA binding of androgen (AR) and glucocorticoid (GR) receptor peptide fragments. To determine its identity, RAF was purified from HeLa cell extracts by anion-exchange and DNA affinity chromatography. RAF activity co-purified with a 110-kDa protein, the partial amino acid sequence of which shares 97.5% identity with insulin degrading enzyme (IDE), a metalloendoprotease implicated in the intracellular degradation of insulin. The identity of RAF was confirmed in gel shift assays by demonstrating that AR.RAF and GR.RAF DNA complexes shifted to a slower mobility in the presence of an IDE antibody. Purified preparations of RAF had insulin degrading activity, and this activity was inhibited by AR. In addition, the interaction of AR or GR with RAF was competed by insulin and bacitracin, a competitive inhibitor of IDE. The interactions of AR and GR with IDE may have important implications for both insulin- and steroid-mediated signaling.

Receptor accessory factor enhances specific DNA binding of androgen and glucocorticoid receptors.

Protein-protein interactions are common among transcriptional activators and may have important consequences for gene regulation. Using the mobility shift assay, we have identified a factor that enhances specific DNA binding of truncated rat androgen (AR) and glucocorticoid (GR) receptors by 25- and 6-fold, respectively, through the formation of heteromeric complexes. This factor, designated receptor accessory factor, or RAF, also potentiates DNA binding of full-length human GR. RAF is temperature and trypsin sensitive and is present in a variety of cultured mammalian cells. By gel filtration RAF has a predicted molecular mass of 130 kDa. RAF enhancement of AR-DNA binding is optimal with androgen response element DNA. RAF appears to interact directly with AR because 1) deoxycholate, which interferes with protein-protein but not protein-DNA interactions, prevents RAF.AR.DNA complex formation, 2) RAF activity is recovered from an androgen response element DNA affinity column only in the presence of AR, and 3) RAF increases the size of an AR.DNA complex by gel filtration. Mutagenesis of truncated AR fragments indicates that a region in the NH2-terminal domain is required for RAF to enhance AR-DNA binding. The interaction of RAF with AR and GR suggests that RAF might influence the ability of these nuclear receptors to activate transcription.

Enhancement of the anabolic effects of growth hormone and insulin-like growth factor I by use of both agents simultaneously.

The use of growth hormone (GH) as an anabolic agent is limited by its tendency to cause hyperglycemia and by its inability to reverse nitrogen wasting in some catabolic conditions. In a previous study comparing the anabolic actions of GH and IGF-I (insulin-like growth factor I), we observed that intravenous infusions of IGF-I (12 micrograms/kg ideal body wt [IBW]/h) attenuated nitrogen wasting to a degree comparable to GH given subcutaneously at a standard dose of 0.05 mg/kg IBW per d. IGF-I, however, had a tendency to cause hypoglycemia. In the present study, we treated seven calorically restricted (20 kcal/kg IBW per d) normal volunteers with a combination of GH and IGF-I (using the same doses as in the previous study) and compared its effects on anabolism and carbohydrate metabolism to treatment with IGF-I alone. The GH/IGF-I combination caused significantly greater nitrogen retention (262 +/- 43 mmol/d, mean +/- SD) compared to IGF-I alone (108 +/- 29 mmol/d; P < 0.001). GH/IGF-I treatment resulted in substantial urinary potassium conservation (34 +/- 3 mmol/d, mean +/- SE; P < 0.001), suggesting that most protein accretion occurred in muscle and connective tissue. GH attenuated the hypoglycemia induced by IGF-I as indicated by fewer hypoglycemic episodes and higher capillary blood glucose concentrations on GH/IGF-I (4.3 +/- 1.0 mmol/liter, mean +/- SD) compared to IGF-I alone (3.8 +/- 0.8 mmol/liter; P < 0.001). IGF-I caused a marked decline in C-peptide (1,165 +/- 341 pmol/liter; mean +/- SD) compared to the GH/IGF-I combination (2,280 +/- 612 pmol/liter; P < 0.001), suggesting maintenance of normal carbohydrate metabolism with the latter regimen. GH/IGF-I produced higher serum IGF-I concentrations (1,854 +/- 708 micrograms/liter; mean +/- SD) compared to IGF-I only treatment (1,092 +/- 503 micrograms/liter; P < 0.001). This observation was associated with increased concentrations of IGF binding protein 3 and acid-labile subunit on GH/IGF-I treatment and decreased concentrations on IGF-I alone. These results suggest that the combination of GH and IGF-I treatment is substantially more anabolic than either IGF-I or GH alone. GH/IGF-I treatment also attenuates the hypoglycemia caused by IGF-I alone. GH/IGF-I treatment could have important applications in diseases associated with catabolism.

Organ perfusion by dynamic scintigraphy convection-diffusion tracer kinetics in a phantom.

Dynamic scintigraphy is used widely to evaluate qualitatively the perfusion of an organ. Attempts to quantify blood flow to an organ by means of scintigraphic imaging modalities have often employed assumptions that lead to oversimplifying the physiology of the tracer kinetics. We used a mathematical formalism described by W. Perl and F. P. Chinard (Circ. Res. 22: 273-298, 1968), the convection-diffusion tracer kinetics, model, for parameter evaluation of flow (F) and volume of distribution (V). This modeling methodology was evaluated using a circulatory phantom with absolute flow measured independently by flowmeter. In a series of 22 phantom experiments with F/V < 0.32 s-1, there was a strong correlation between F and flow probe measurement [r = 0.97; slope = 1.08 +/- 0.06 (SE)]. The theoretical analysis comparing this approach with classical tracer kinetics methods explains both the satisfactory results for F/V using mean transit time and the systematic overestimation of F/V using decay constant methods.