A MEMS nanoindenter with an integrated AFM cantilever gripper for nanomechanical characterization of compliant materials.

This work presents the development of a MEMS nanoindenter that uses exchangeable AFM probes for quasi-static nanomechanical characterization of compliant and ultra-compliant materials. While the electrostatic micro-force transducer of the MEMS nanoindenter provides a maximum indentation depth up to 9.5 µm with a maximum output force of 600 µN, experimental investigations reveal that it can achieve a depth and force resolution better than 4 pm Hz-1/2 and 0.3 nN Hz-1/2, in air for f≥ 1 Hz. A passive AFM probe gripper is integrated into the MEMS nanoindenter, allowing the nanoindenter to utilize various AFM probes as an indenter for material testing. A proof-of-principle experimental setup has been built to investigate the performance of the MEMS nanoindenter prototype. In proof-of-principle experiments, the prototype with a clamped diamond AFM probe successfully identified an atomic step (∼0.31 nm) within a Si < 111 > ultraflat sample using the scanning probe microscopy mode. The nanomechanical measurement capability of the MEMS nanoindenter prototype has been verified by means of measurements of reference polymer samples using a silicon AFM probe and by means of measurements of the elastic properties of a PDMS sample using a spherical diamond-coated AFM probe. Owing to its compact and low-cost but high-resolution capacitive readout system, this MEMS nanoindenter head can be further applied for in-situ quantitative nanomechanical measurements in AFMs and SEMs.

Note: Nanomechanical characterization of soft materials using a micro-machined nanoforce transducer with an FIB-made pyramidal tip.

The quantitative nanomechanical characterization of soft materials using the nanoindentation tech-nique requires further improvements in the performances of instruments, including their force resolution in particular. A micro-machined silicon nanoforce transducer based upon electrostatic comb drives featuring the force and depth resolutions down to ∼1 nN and 0.2 nm, respectively, is described. At the end of the MEMS transducer's main shaft, a pyramidal tip is fabricated using a focused ion beam facility. A proof-of-principle setup with this MEMS nanoindenter has been established to measure the mechanical properties of soft polydimethylsiloxane. First measurement results demonstrate that the prototype measurement system is able to quantitatively characterize soft materials with elastic moduli down to a few MPa.

Functional modeling of cobalamine-independent methionine synthase with pyrazolylborate-zinc-thiolate complexes.

A series of new pyrazolylborate-zinc-thiolate complexes Tp(Ph,Me)Zn-SR and Tp(Me,Me)Zn-SR, including two homocysteine derivatives, were prepared and structurally characterized. Their reactions with methyl iodide in nonpolar media resulted in the formation of the thioethers MeSR, including two methionine derivatives, and Tp(R',Me)Zn-I in all cases. Methylation of the thiolates could also be achieved with dimethyl sulfate and trimethylsulfonium iodide but not with trimethyl phosphate or N-methylpyridinium salts. The accumulated evidence indicates that the methylation occurs intramolecularly, i.e., at the zinc-bound thiolates: (i) The reactions occur readily in nonpolar media. (ii) Thiolate exchange at Tp(Ph,Me)Zn-SR with [PPN]SR' is slower than thiolate alkylation. (iii) The methylation of Tp(Ph,Me)Zn-SBn with MeI is a clean second-order reaction with k'' = 1.75 x 10(-2) M(-1) s(-1) at 300 K.

Synthesis and characterization of the dimercury(I)-linked compound [PPn]4[(Re7C(CO)21Hg)2]. Oxidative cleavage of the mercury-mercury bond leading to carbidoheptarhenate complexes of mercury(II), including [PPN][Re7C(CO)21Hg(S=C(NME2)2)].

The reaction of [PPN](3)[Re(7)C(CO)(21)] with Hg(2)(NO(3))(2).2H(2)O in dichloromethane formed the complex [PPN](4)[(Re(7)C(CO)(21)Hg)(2)] ([PPN](4)[1]), isolated in 60% yield. Analogous salts of [1](4-) with [PPh(4)](+) and [NEt(4)](+) were also prepared. The crystal structure of [PPN](4)[1] showed that two carbidoheptarhenate cores are linked by a dimercury(I) unit (d(Hg-Hg) = 2.610(4) A), with each individual mercury atom face-bridging. Oxidative cleavage of the Hg-Hg bond in [1](4-) was effected by 4-bromophenyl disulfide to form [Re(7)C(CO)(21)HgSC(6)H(4)Br](2-) ([4](2-)), by I(2) to form [Re(7)C(CO)(21)HgI](2-) ([5](2-)), and by Br(2) to form [Re(7)C(CO)(21)HgBr](2-) ([6](2-)). Oxidation of [1](4-) by ferrocenium ion (2 equiv) in the presence of tetramethylthiourea resulted in the derivative [Re(7)C(CO)(21)HgSC(NMe(2))(2)](-) ([7](-)). The molecular structure of [PPN][7] was determined by X-ray crystallography. This is the first example of a carbidoheptarhenate-mercury complex with a neutral ligand on mercury, and ligand exchange was demonstrated by displacement with triethylphosphine. Complex [7](-) can also be prepared by protonating [Re(7)C(CO)(21)HgO(2)CCH(3)](2-) in the presence of tetramethylthiourea. Cyclic voltammetry data to calibrate and compare the redox properties of compounds [1](4-) and [7](-) have been measured.

Inhibition of human allergic T-cell responses by IL-10-treated dendritic cells: differences from hydrocortisone-treated dendritic cells.

BACKGROUND: Dendritic cells (DCs) are able to induce human allergic T(H)1 responses as well as T(H)2 responses. OBJECTIVE: In this study, we examined the effect of antiinflammatory agents such as IL-10 and hydrocortisone (HC) on the accessory function of DCs and the resulting T-cell response, especially that of T(H)2 cells. METHODS: Naive and memory CD4(+) T cells from atopic donors were stimulated with autologous allergen-pulsed DCs generated from CD14(+) monocytes by culture with GM-CSF/IL-4 and fully matured with IL-1 beta, TNF-alpha, and PGE(2) in the presence or absence of IL-10 or HC. RESULTS: IL-10-treated DCs and, to a lesser extent, HC-treated DCs showed a decreased expression of MHC II molecules, the costimulatory molecule CD86, and the DC-specific marker CD83, as well as a strongly reduced IL-12 secretion. Consequently, T-cell proliferation was reduced after stimulation with IL-10- or HC-treated DCs alike. However, pretreatment of DCs with IL-10 inhibited the production of T(H)1 and T(H)2 cytokines by T cells, whereas HC-treated DCs inhibited production of IFN-gamma but induced an increased release of IL-4 and no change in IL-5. Both effects were long-lasting; cytokine production remained low (which was due not to enhanced apoptosis but to functional hyporesponsiveness) or even increased after restimulation with fully matured DCs. CONCLUSION: These data indicate that IL-10- or HC-treated DCs differ in their ability to influence human allergic T-cell responses. This has major implications for therapeutic strategies aiming at the downregulation of proallergic T(H)2 responses.

Control of cell fate in plant meristems.

In contrast to animals, plants do not stop to initiate organs with the end of embryogenesis. Instead, most of the growth and development of higher plants will take place during later phases following germination of the seed. Plant development depends on the activity of two meristems, the root meristem and the shoot apical meristem (SAM), that are located at opposite ends of the plant embryo. These meristems serve as a source of pluripotent stem cells and, in case of the SAM, provide a centre for repetitive organ initiation. In order to maintain the SAM throughout plant life, the cells that are lost from the meristem through organ initiation and differentiation have to be replaced from the stem cell population. In this paper, we will discuss recent results indicating that the fate of stem cells in plant meristems is controlled by directional signalling systems.

Autoimmune hemolytic anemia caused by IgG lambda-monotypic cold agglutinins of anti-Pr specificity after rubella infection.

BACKGROUND: In postinfection cold agglutinin (CA) disease, a relation between CA specificity and the underlying infectious agent has been observed. The induction of anti-I by Mycoplasma pneumoniae and that of anti-i by EBV are well-established examples. CASE REPORT: A 5-year-old boy developed severe hemolytic anemia after serologically ascertained rubella infection. Hemolysis was caused by high-titer CAs, which were analyzed by absorption and elution with sialidase-treated RBCs and hemagglutination-inhibition experiments. RESULTS: After elimination of normal anti-I and anti-T, the predominant CA was found to be an IgG lambda autoantibody with anti-Pr(1) specificity. CONCLUSION: This case seems to be of interest because it is the first report of severe CA-induced hemolysis after rubella infection, it is the first description of an IgG lambda-monotypic CA, and, along with previous case reports (three established and three suspected cases), it indicates a relationship between rubella infection and the CA specificity anti-PR:

Functional domains in plant shoot meristems.

The development of higher plants depends on the activity of a shoot apical meristem. Organs are formed on the flanks of the meristem, while pluripotent stem cells are found in a separate domain in the meristem centre. Further domains are distinguished by the expression patterns of genes that control the development of the shoot meristem. Although most plant cells are immobile, their relative position within a meristem, and therefore also their function, can change after cell divisions. To maintain an active shoot meristem throughout plant life, the cells in the meristem need constantly to assess their position, transmit this information to others, and readjust their gene expression profiles and their fate. Some of the genes that permit intercellular communication have been isolated. They enable the flow of information in and between meristem regions via ligands and receptor proteins to transcription factors, that ultimately control the fate of cells in the centre of the meristem.

Mental health care in Germany: carers' perspectives.

OBJECTIVE: To assess the mental health care system in Germany from the point of view of the federal association of family carers of people with mental illness. METHOD: Family carer involvement and perspective are discussed on the basis of available literature, questionnaire surveys and documents of carer organizations. RESULTS: At the beginning of the reform movement the views of informal carers were not discussed. Since 1985 family carers have joined forces to express their views on needs of the severely mentally ill and their carers. Their aim is to point out deficits of the care system and to work towards improved care for their relatives with mental illness and changes in the mental health care system. CONCLUSION: In the reform process informal carers should receive support and be respected as experts and partners.

European perspectives: a carer's view.

OBJECTIVE: To present the work of the European Federation of Associations of Families of Mentally III People (EUFAMI) and discuss issues of concern to family carers. METHOD: The problem areas identified and discussed by family carers are presented on the basis of questionnaire surveys organized by EUFAMI. Addresses of national organisations of family carers are included. RESULTS: A range of problem areas are identified; they include subsistence and welfare payments for the severely mentally ill, some shortage of general hospital units, problems of care co-ordination, issues of respect for family carers and family involvement. CONCLUSION: The aim of best practice in mental health care throughout Europe has not yet been reached. Key activities of EUFAMI are aimed at empowerment of families and best practice in psychiatry in Europe.

Dependence of stem cell fate in Arabidopsis on a feedback loop regulated by CLV3 activity.

The fate of stem cells in plant meristems is governed by directional signaling systems that are regulated by negative feedback. In Arabidopsis thaliana, the CLAVATA (CLV) genes encode the essential components of a negative, stem cell-restricting pathway. We used transgenic plants overexpressing CLV3 to show that meristem cell accumulation and fate depends directly on the level of CLV3 activity and that CLV3 signaling occurs exclusively through a CLV1/CLV2 receptor kinase complex. We also demonstrate that the CLV pathway acts by repressing the activity of the transcription factor WUSCHEL, an element of the positive, stem cell-promoting pathway.

Comparison of allergen-stimulated dendritic cells from atopic and nonatopic donors dissecting their effect on autologous naive and memory T helper cells of such donors.

BACKGROUND: Because of their production of IL-12, mature dendritic cells (DC) are potent inducers of T(H)1 responses. However, recent reports have demonstrated that DCs can also induce T(H)2 differentiation. OBJECTIVE: In the current study we investigated which immune response is induced by DCs in naive CD45RA(+) or memory CD45R0(+) CD4(+) T cells from atopic individuals (patients with grass pollen, birch pollen, or house dust mite allergy) compared with nonatopic control subjects. METHODS: Immature DCs, generated from peripheral blood monocytes from atopic and nonatopic donors, were pulsed with the respective allergen and fully matured. Then the mature DCs were cocultured in vitro with autologous naive (CD45RA(+)) and memory (CD45R0(+)) CD4(+) T cells and cytokine and IgE production were measured by ELISA. RESULTS: After the second restimulation with allergen-pulsed DCs, naive as well as memory autologous CD4(+) T cells from atopic but not from nonatopic donors showed an enhanced production of the T(H)2-type cytokines IL-4, IL-5, and IL-10, resulting in an increased IgE production, whereas IFN-gamma production and proliferation were not different. IL-12 production and surface marker expression of DCs derived from atopic and nonatopic donors did not differ and addition of neutralizing anti-IL-12 mAbs did not increase IL-4 but diminished IFN-gamma production. CONCLUSION: These data indicate that mature DCs are able to induce naive and activate allergen-specific T helper cells to produce T(H)2 cytokines if the T cells are derived from atopic donors. This phenomenon is not due to diminished IL-12 production by DCs of atopic donors.

Carbidohexarhenate cluster cores bicapped by mercury with acetate or thiolate ligands.

The reaction of [PPh4]3[Re7C(CO)21] (1) with 1 or more equiv of Hg(OAc)2 in dichloromethane provides the monomercury derivative [PPh4]2[Re7C(CO)21HgOAc] (2) in high yield. However, in the presence of methanol the reaction of 1 with 2 equiv of Hg(OAc)2 yields the dimercury hexarhenium cluster compound [PPh4]2[Re6C(CO)18(HgOAc)2] (3) together with the dirhenium complex [PPh4][Re2(CO)6(mu-OMe)2(mu-OAc)] (4). The dimercury compound 3 reacts with various thiols HS-Z to form thiolate-substituted derivatives [PPh4]2[Re6C(CO)18(HgSZ)2] [Z = C6H4Br (5); C5H4N (6); C2H4COOH (7)]. All new compounds have been characterized by a combination of analytical and spectroscopic data, and the molecular structures of compounds 3-6 have been determined by X-ray crystallography.

Allergen-specific immune deviation from a TH2 to a TH1 response induced by dendritic cells and collagen type I.

BACKGROUND: Atopy and IgE production are associated with enhanced allergen-specific T(H)2 responses. Therefore a causative treatment may result from the deviation of this T(H)2-dominated immune response toward a T(H)1 response. OBJECTIVE: This study was carried out to analyze whether dendritic cells, the most potent antigen-presenting cells that are also known to induce antigen-specific T(H)1 responses, are suitable for therapy of atopic diseases by shifting the allergen-specific T(H)2 response toward a T(H)1 response. METHODS: Monocyte-derived dendritic cells were used to present allergens in vitro to autologous CD4(+) T cells of allergic persons. Because collagen type I activates dendritic cells and enhances the secretion of IL-12, we performed allergen presentation assays also in the presence of collagen type I. RESULTS: After stimulation with allergen-pulsed dendritic cells the production of IFN-gamma as well as that of IL-4 and IL-5 by CD4(+) T cells was enhanced. In the presence of collagen type I, however, a significant shift toward a T(H)1 response with increased production of IFN-gamma and a decreased production of IL-5 could be observed. When T cells were stimulated directly with anti-CD3 and anti-CD28 in the absence of antigen-presenting cells, it was demonstrated that collagen type I also exerted a direct effect on T cells, increasing their IFN-gamma production. CONCLUSION: These data indicate that collagen type I influences dendritic cells as well as T cells in a way that a shift in cytokine production results in a T(H)1 response even in already-sensitized atopic individuals.

Signaling of cell fate decisions by CLAVATA3 in Arabidopsis shoot meristems.

In higher plants, organogenesis occurs continuously from self-renewing apical meristems. Arabidopsis thaliana plants with loss-of-function mutations in the CLAVATA (CLV1, 2, and 3) genes have enlarged meristems and generate extra floral organs. Genetic analysis indicates that CLV1, which encodes a receptor kinase, acts with CLV3 to control the balance between meristem cell proliferation and differentiation. CLV3 encodes a small, predicted extracellular protein. CLV3 acts nonautonomously in meristems and is expressed at the meristem surface overlying the CLV1 domain. These proteins may act as a ligand-receptor pair in a signal transduction pathway, coordinating growth between adjacent meristematic regions.

Signals involved in the early TH1/TH2 polarization of an immune response depending on the type of antigen.

BACKGROUND: The early production of distinct cytokines by epidermal cells (ECs) in response to antigen exposure may govern the development of TH1 -like immune responses, such as contact sensitivity, or TH2 -like immune responses, such as IgE-dependent allergies of the immediate type, depending on the type of antigen. OBJECTIVE: The aim of this study was to compare the signals induced by protein allergens with those induced by haptens in ECs and subsequently in local draining lymph node cells (LNCs) or splenocytes. METHODS: BALB/c mice were primed in vivo with the protein allergens ovalbumin or birch pollen or the haptens 2, 4-dinitrofluorobenzene or trinitrochlorbenzene, respectively, and cytokine and immunoglobulin secretions of responding splenocytes were measured by ELISA after in vitro coculture with ECs. Induction of cytokine mRNA expression in ECs and LNCs was analyzed by reverse transcriptase-PCR. RESULTS: In the presence of protein allergens, ECs enhance the induction of a TH2 immune response (IL-4 and IgE production of splenocytes), whereas a TH1 immune response (IFN-gamma and IgG2a production) was only induced in the context of haptens. Heat inactivation of ovalbumin did not diminish the development of a TH2 immune response. One direct effect of antigen on ECs was the earlier expression of IL-10 mRNA after stimulation with protein allergens (30 minutes) than with haptens (2 hours) in vitro. By using an in vivo approach, sensitization of the skin with trinitrochlorbenzene, but not with ovalbumin, resulted in an early induction of IL-1beta, IL-12p40, and IFN-gamma mRNA in LNCs, whereas IL-18 was induced by both. CONCLUSION: These data indicate that the type of antigen strongly influences the type of immune response by eliciting distinct signals already in the epithelium.

Influence of extracellular matrix proteins on the development of cultured human dendritic cells.

The development of dendritic cells (DC) is still only partly understood. Recently established culture systems using CD34+ cells or monocytes as precursor cells for the generation of DC indicate the necessity of pro-inflammatory cytokines for their development. In vivo the contact to other cells or to the proteins of the extracellular matrix might also be essential for their development. In our experiments we used granulocyte-macrophage colony-stimulating factor- and IL-4-treated human monocytes as precursor cells to investigate the interaction of DC at different maturation stages with the matrix proteins fibronectin, collagen type I and collagen type IV. We demonstrate a strong beta1-integrin-mediated adherence of immature DC to fibronectin that is lost completely during maturation. The binding to collagen type I was less strong but induced a maturation of the precursor cells. After 3 days of culture on this protein, the cells showed all features of fully matured DC such as expression of CD83 and an excellent allostimulatory capacity. The reason for this effect was shown to be the induction of TNF-alpha production by the DC themselves. In contrast to the adhesion to fibronectin, the maturation and the cytokine production of DC induced by collagen type I could not be inhibited by blocking of beta1-integrins. These results indicate that proteins of the extracellular matrix play an important role in the development and function of human DC.

[Food hygiene aspects in the production of food fish in fishing]. / Lebensmittelhygienische Aspekte bei der Gewinnung von Speisefischen in der Angelfischerei.

The development of the aerob-mesophilic bacteria on epidermis and peritoneum of 68 barbels was determined at 0, 4 and 8 hours after slaughtering. Therefore, one group of 34 animals was stored at 15.3 degrees C, an other equal one at 21.6 degrees C. A change in germ counts per cm2 could be seen in none of the groups during the first 4 hours. However, unrefrigerated carcasses showed an increase of bacteria up to 5-fold between the 4th and 8th hour, whereas in the refrigerated group no change occurred during that time, too. Rinsing the fish after slaughtering resulted in a decrease of the initial bacterial counts by up to 65.4% and so in significantly lower germ loads at the end of the storage time. These results were confirmed by contaminating 24 rainbow trout with Salmonella Infantis artificially. The frequency of detection did not change in refrigerated fish over 8 hours, while nearly doubling in unrefrigerated ones. Moreover, it could be shown that a Salmonella-concentration of only 30 CFU per 100 ml water was sufficient for contaminating fish in detectable grades. The study leads to the conclusion that the storage of instantly slaughtered fish in a common thermobox with freezing elements is suited for preserving its microbiological status for at least 8 hours. The caging of living fish after capture, which must be regarded critically under the aspect of treating animals in a humane way, seems therefore unnecessary.

[Severe neonatal alloimmune thrombocytopenia with delayed antibody detection]. / Schwere neonatale Alloimmunthrombozytopenie mit verzögertem Antikörpernachweis.

Neonatal alloimmune thrombocytopenia (NAIT) is caused by maternal immunisation against a paternal antigen on fetal platelets. The antigen involved in the majority of cases is HPA-1 a (PIA1). Usually circulating platelet alloantibodies are detectable in the mother. In this report, we present a thrombocytopenic newborn with severe hemorrhagic diathesis due to materno-fetal HPA-1 a (PIA1) incompatibility. Platelet antibodies could initially not be demonstrated in the mother's serum but became detectable after four weeks. Because of the severe and protracted course of the disease, repeated platelet substitution was necessary throughout the first two months of life.