[Chimeric antigen receptors (CARs): universal tools in adoptive cell therapy]. / Chimäre Antigenrezeptoren (CAR) ­ universelle Werkzeuge in der zellulären Immuntherapie.

BACKGROUND: The observation that tumor-infiltrating lymphocytes (TIL) after ex vivo amplification can control tumors in the long term led to the concept of redirecting patients' cytolytic T­cells by a receptor with defined specificity against the tumor. OBJECTIVES: Development of a recombinant receptor-signal molecule (chimeric antigen receptor, CAR) to increase selectivity and enhance anti-tumor immunity. METHODS: Description of a prototype CAR, overview of the modular composition and further development of CAR technology for use in adoptive immune cell therapy. RESULTS: Intensive research over the last two decades has shown how CAR-mediated T­cell activation is influenced by factors such as binding affinity, the epitope of the target antigen, its expression density and accessibility on the tumor cells, as well as by the signaling domains and their combination to induce T­cell activation. The quality and duration of the T­cell response can be specifically modulated by modifying the modular composition of the CAR; CAR T­cells can act as "biopharmaceutical factories" (T-cells redirected for unrestricted cytokine-mediated killing, TRUCK) in the tissue by CAR-mediated release of transgenic therapeutic proteins. CONCLUSION: Adoptive CAR T­cell therapy has shown clinical efficacy in the treatment of hematological malignancies; the treatment of solid tumors, however, is more challenging. Allogeneic CAR T­cell technology is aimed at generating "off-the-shelf" CAR T­cells that are accessible for a large number of patients. A further promising approach is the use of CAR T­cells for other therapeutic applications such as the treatment of autoimmune diseases.

A new microscopic method to analyse desiccation-induced volume changes in aeroterrestrial green algae.

Aeroterrestrial green algae are exposed to desiccation in their natural habitat, but their actual volume changes have not been investigated. Here, we measure the relative volume reduction (RVRED ) in Klebsormidium crenulatum and Zygnema sp. under different preset relative air humidities (RH). A new chamber allows monitoring RH during light microscopic observation of the desiccation process. The RHs were set in the range of ∼4 % to ∼95% in 10 steps. RVRED caused by the desiccation process was determined after full acclimation to the respective RHs. In K. crenulatum, RVRED (mean ± SE) was 46.4 ± 1.9%, in Zygnema sp. RVRED was only 34.3 ± 2.4% at the highest RH (∼95%) tested. This indicates a more pronounced water loss at higher RHs in K. crenulatum versus Zygnema sp. By contrast, at the lowest RH (∼4%) tested, RVRED ranged from 75.9 ± 2.7% in K. crenulatum to 83.9 ± 2.2% in Zygnema sp. The final volume reduction is therefore more drastic in Zygnema sp. These data contribute to our understanding of the desiccation process in streptophytic green algae, which are considered the closest ancestors of land plants.

Photosynthetic efficiency, desiccation tolerance and ultrastructure in two phylogenetically distinct strains of alpine Zygnema sp. (Zygnematophyceae, Streptophyta): role of pre-akinete formation.

Two newly isolated strains of green algae from alpine regions were compared physiologically at different culture ages (1, 6, 9 and 15 months). The strains of Zygnema sp. were from different altitudes ('Saalach' (S), 440 m above sea level (a.s.l.), SAG 2419 and 'Elmau-Alm' (E-A), 1,500 m a.s.l., SAG 2418). Phylogenetic analysis of rbcL sequences grouped the strains into different major subclades of the genus. The mean diameters of the cells were 23.2 µm (Zygnema S) and 18.7 µm (Zygnema E-A) but were reduced significantly with culture age. The photophysiological response between the strains differed significantly; Zygnema S had a maximal relative electron transport rate (rETR max) of 103.4 µmol electrons m(-2) s(-1), Zygnema E-A only 61.7 µmol electrons m(-2) s(-1), and decreased significantly with culture age. Both strains showed a low-light adaption and the absence of strong photoinhibition up to 2,000 µmol photons m(-2) s(-1). Photosynthetic oxygen production showed similar results (P max Zygnema S, 527.2 µmol O2 h(-1) mg(-1) chlorophyll (chl.) a, Zygnema E-A, 390.7 µmol O2 h(-1) mg(-1) chl. a); the temperature optimum was at 35 °C for Zygnema S and 30 °C for Zygnema E-A. Increasing culture age moreover leads to the formation of pre-akinetes, which accumulate storage products as revealed by light and transmission electron microscopy. Desiccation at 84 % relative air humidity (RH) lead to a reduction of the effective quantum yield of photosystem II (PSII) (ΔFv/Fm') to zero between 90 to 120 min (Zygnema S) and between 30 to 60 min (Zygnema E-A), depending on the culture age. A partial recovery of ΔFv/Fm' was only observed in older cultures. We conclude that pre-akinetes are crucial for the aeroterrestrial lifestyle of Zygnema.

The weal and woe of costimulation in the adoptive therapy of cancer with chimeric antigen receptor (CAR)-redirected T cells.

Adoptive cell therapy has shown impressive efficacy to combat cancer in early phase clinical trials, in particular when T cells engineered to specifically target tumor cells were applied. The patient's T cells are genetically equipped with a chimeric antigen receptor (CAR) which allows them to be redirected in a predefined manner towards virtually any target; by using an antibody-derived domain for binding, CAR T cells can be redirected in a major histocompatibility complex (MHC) dependent and independent fashion. The CAR also provides the stimuli required to induce and maintain T cell activation. Recent clinical data sustain the notion that strong costimulation in conjunction with the primary activation signal is crucial for lasting therapeutic efficacy of CAR T cells. However, costimulation is a double-edged sword and the impact of the individual costimuli to optimize T cell activation is still under debate; some general rules are emerging. The review summarizes how costimulation modulates, improves and prolongs the redirected anti-tumor T cell response and how the same costimulatory signals may contribute to unintended side effects including "cytokine storm" and T cell repression. Upcoming strategies to break the activation/repression circle by using CAR's with modified costimulatory signals are also discussed.

Psychiatric services in the community? The German public's opinion in 1990 and 2011.

Background. During the last two decades, the change from custodial care provided by large institutions to community-focused services made considerable progress in Germany. However, nothing is known about how this is reflected in the public's acceptance of community psychiatry services. Methods. The study is based on data from two population surveys among German citizens aged 18 years and over, living in the 'old' German States. The first was conducted in 1990 (n = 3067), the second in 2011 (n = 2416). With the help of identical questions, respondents' attitudes towards psychiatric units at general hospitals and group homes for mentally ill people were assessed. Results. While the proportion of the public that explicitly welcomed establishing psychiatric units at general hospitals and opening group homes for mentally ill people decreased, the proportion of those who reacted with indifference increased. The proportion of the German population that explicitly rejected the implementation of these services remained unchanged. Conclusions. While community psychiatry services expanded considerably over the last few years, the public's attitude towards them has not changed substantially.

Spherical particles of halophilic archaea correlate with exposure to low water activity--implications for microbial survival in fluid inclusions of ancient halite.

Viable extremely halophilic archaea (haloarchaea) have been isolated from million-year-old salt deposits around the world; however, an explanation of their supposed longevity remains a fundamental challenge. Recently small roundish particles in fluid inclusions of 22 000- to 34 000-year-old halite were identified as haloarchaea capable of proliferation (Schubert BA, Lowenstein TK, Timofeeff MN, Parker MA, 2010, Environmental Microbiology, 12, 440-454). Searching for a method to produce such particles in the laboratory, we exposed rod-shaped cells of Halobacterium species to reduced external water activity (a(w)). Gradual formation of spheres of about 0.4 µm diameter occurred in 4 M NaCl buffer of a(w) ≤ 0.75, but exposure to buffered 4 M LiCl (a(w) ≤ 0.73) split cells into spheres within seconds, with concomitant release of several proteins. From one rod, three or four spheres emerged, which re-grew to normal rods in nutrient media. Biochemical properties of rods and spheres were similar, except for a markedly reduced ATP content (about 50-fold) and an increased lag phase of spheres, as is known from dormant bacteria. The presence of viable particles of similar sizes in ancient fluid inclusions suggested that spheres might represent dormant states of haloarchaea. The easy production of spheres by lowering a(w) should facilitate their investigation and could help to understand the mechanisms for microbial survival over geological times.

Gender differences in public beliefs and attitudes about mental disorder in western countries: a systematic review of population studies.

AIMS: While quite a number of theories and hypotheses about gender differences in public beliefs and attitudes about mental illness have been proposed, the empirical evidence, particularly evidence based on population studies, is rather scarce. METHODS: A systematic review of population-based studies providing information on gender differences in beliefs about mental disorders and attitudes towards the mentally ill was carried out. RESULTS: While both genders are no different in their willingness to seek informal help for mental illness, women seem more ready to recommend professional help than men. They also evaluate treatment outcomes more favourably. Women are more likely to endorse psychosocial conceptualizations of mental illness than men, and, in consequence, are more in favour of psychotherapy. With a few exceptions, women do not seem to display more favourable attitudes than men towards people with mental disorder. Female patients seem to be rejected by the public less than male patients. CONCLUSIONS: Our review suggests that gender matters in public beliefs and attitudes about mental illness. Some theoretical assumptions are supported by empirical findings, others not. However, as evidence is rather scarce, further studies testing theory-driven hypotheses are needed.

Evolution of public attitudes about mental illness: a systematic review and meta-analysis.

OBJECTIVE: To explore whether the increase in knowledge about the biological correlates of mental disorders over the last decades has translated into improved public understanding of mental illness, increased readiness to seek mental health care and more tolerant attitudes towards mentally ill persons. METHOD: A systematic review of all studies on mental illness-related beliefs and attitudes in the general population published before 31 March 2011, examining the time trends of attitudes with a follow-up interval of at least 2 years and using national representative population samples. A subsample of methodologically homogeneous studies was further included in a meta-regression analysis of time trends. RESULTS: Thirty-three reports on 16 studies on national time trends met our inclusion criteria, six of which were eligible for a meta-regression analysis. Two major trends emerged: there was a coherent trend to greater mental health literacy, in particular towards a biological model of mental illness, and greater acceptance of professional help for mental health problems. In contrast, however, no changes or even changes to the worse were observed regarding the attitudes towards people with mental illness. CONCLUSION: Increasing public understanding of the biological correlates of mental illness seems not to result in better social acceptance of persons with mental illness.

Sequential targeting of CFTR by BAC vectors generates a novel pig model of cystic fibrosis.

Cystic fibrosis (CF) is the most common lethal inherited disease in Caucasians and is caused by mutations in the CFTR gene. The disease is incurable and medical treatment is limited to the amelioration of symptoms or secondary complications. A comprehensive understanding of the disease mechanisms and the development of novel treatment options require appropriate animal models. Existing CF mouse models fail to reflect important aspects of human CF. We thus generated a CF pig model by inactivating the CFTR gene in primary porcine cells by sequential targeting using modified bacterial artificial chromosome vectors. These cells were then used to generate homozygous CFTR mutant piglets by somatic cell nuclear transfer. The homozygous CFTR mutants lack CFTR protein expression and display severe malformations in the intestine, respiratory tract, pancreas, liver, gallbladder, and male reproductive tract. These phenotypic abnormalities closely resemble both the human CF pathology as well as alterations observed in a recently published CF pig model which was generated by a different gene targeting strategy. Our new CF pig model underlines the value of the CFTR-deficient pig for gaining new insight into the disease mechanisms of CF and for the development and evaluation of new therapeutic strategies. This model will furthermore increase the availability of CF pigs to the scientific community.

What to do about depression? Help-seeking and treatment recommendations of the public.

AIMS: Several population studies on beliefs about depression carried out in western countries during the 1990s have shown that the public clearly favors psychotherapy over antidepressant medication. The present study examines whether this phenomenon still exists at the end of the first decade of the twenty-first century. MATERIALS AND METHODS: In 2009, a telephone survey was conducted among the population of Vienna aged 16 years and older (n = 1205). A fully structured interview was administered which began with the presentation of a vignette depicting a case of depression fulfilling the diagnostic criteria of DSM-IV for a moderate depressive episode. RESULTS: Psychotherapists were most frequently endorsed as source of professional help. Antidepressant medication still was more frequently advised against than recommended. Respondents familiar with the treatment of depression tended to be more ready to recommend to seek help from mental health professionals and to endorse various treatment options, particularly medication. CONCLUSION: At the end of the first decade of this century, there still exists a large gap between the public's beliefs and what mental health professionals consider appropriate for the treatment of depression. Therefore, further effort to improve the public's mental health literacy seems necessary.

The loss of sadness: the public's view.

OBJECTIVE: With the exception of bereavement, the diagnosis of major depressive disorder in the DSM-IV does not take into account the context in which the symptoms occur. Recent criticism has maintained that common sense suggests making a distinction between depression as mental disorder and sorrow as 'normal' reaction to social stress. This study sets out to investigate whether the public does in fact make this distinction. METHOD: In spring 2009, a population-based survey was conducted by phone in the city of Vienna (n = 1205). A fully structured interview was carried out which began with the presentation of a vignette describing a diagnostically unlabeled case of depression, with or without provision of information about preceding stressful life events. RESULTS: Respondents presented with vignettes containing information on loss events were less likely to define depressive symptoms as indication of mental illness. They were also not as willing to recommend professional help. And if they were, they tended to less frequently recommend seeking help from someone with a medical background. CONCLUSION: In contrast to the conceptualization in the DSM-IV, the public tends to perceive depressive symptoms differently depending on the context in which they occur.

Cell morphology, ultrastructure, and calcification pattern of Oocardium stratum, a peculiar lotic desmid.

Cell morphology and ultrastructure of the desmid Oocardium stratum and its habitat conditions in two limestone-precipitating spring habitats in the Alps were studied. In spite of specific cell geometry, we found ultrastructural features (nucleus with nucleolus, Golgi apparatus, chloroplast structure, lipid bodies, cell wall texture) closely related to other desmids. The type of the mucilage pore apparatus perforating in high densities extended areas of the cell wall of Oocardium is of the Cosmarium type. Oocardium contrasts to Cosmarium by a peculiar bilateral cell geometry (lateral sphenoid shape) which is combined with a dislocated nucleus. Although the cell features of Oocardium did not differ between the two habitats, different calcification types (rhombohedral calcite versus fascicular-fibrous calcite) and calcification intensities were recorded. The spatial positioning and extension of the Oocardium niches differed considerably between the two springs in spite of high CO(2) oversaturation at both sites.

Cytoarchitecture of the desiccation-tolerant green alga Zygogonium ericetorum.

In this study, the filamentous green alga Zygogonium ericetorum (Zygnematales, Chlorophyta), collected at its natural habitat in the high alps, was investigated by light, scanning, and transmission electron microscopy. The field samples were separated into a moist fraction when wetted by splattering water of a nearby spring or a desiccated one when visually dried out. Light microscopy demonstrated a purple pigmentation of the sun-exposed upper layers, the central position of the nucleus, and the starch content in the pyrenoids. The smooth surface of the cells occasionally covered with fungal hyphae was shown by scanning electron microscopy. The cytoarchitecture of moist cells revealed many vacuoles and only a thin cytoplasmic area surrounding the two chloroplasts. The secondary cell walls of older cells were up to 4 microm thick. Organelle membranes as well as thylakoid membranes occasionally showed an inversion of contrast. In the chloroplasts, distinct areas with granular content surrounding the pyrenoids were detected. Within the cytoplasm, electron-dense particles with electron-translucent crystalloid structures were observed. When desiccated samples were investigated, the vacuoles and cytoplasmatic portions appeared destroyed, whereas nucleus and chloroplasts generally remained intact. The thylakoid membranes of desiccated samples showed lumen dilatations and numerous plastoglobules. Water-soluble extracts were separated by high-pressure liquid chromatography that revealed two major compounds with UV-absorbing capacities.

Congenital Central Hypoventilation Syndrome due to PHOX2b gene defects: inheritance from asymptomatic parents.

BACKGROUND: Congenital Central Hypoventilation Syndrome (CCHS, Ondine's curse) is a rare syndrome of dysfunction of the autonomic nervous system characterized by a decreased response to hypercarbia requiring mechanical ventilation in most cases. CCHS is an autosomal-dominant disease associated with tumors of neural crest origin, segmental aganglionosis of the colon, and diffuse autonomic dysregulation symptoms. Most cases of CCHS are caused by de novo heterozygous in-frame expansions within in the PHOX2b gene. PATIENTS AND MAIN RESULTS: Here we report two families in which a PHOX2b defect was inherited from an asymptomatic parent. In family 1 an asymptomatic mother carried a mild mutation (15 bp expansion within the polyalanine repeat) also found in her daughter who was symptomatic immediately after birth but did not require mechanical ventilation. In family 2, two newborn infants with respiratory failure due to insufficient respiratory drive requiring mechanical ventilation were born to asymptomatic parents. A 39 pb expansion within the PHOX2b polyalanine repeat was found in one patient in whom DNA was available, but not in blood leukocytes from any parent. Microsatellite analyses confirmed the identity of the parents, such that a germline mosaicism has to be deduced. CONCLUSIONS: Carriers of mild PHOX2b mutations causing disease in their offspring may be asymptomatic; Modifier genes determining the clinical course may exist. Germline mosaicism may lead to CCHS in children from unaffected parents. Genetic counseling should include these variations.

Mental health literacy and attitude towards people with mental illness: a trend analysis based on population surveys in the eastern part of Germany.

BACKGROUND: There is growing evidence that mental health literacy has improved in western countries in recent years. The question arises as to whether this trend is paralleled by an improvement of attitudes towards people with mental illness. AIM: To examine the development of mental health literacy and the desire for social distance towards people with schizophrenia and major depressive disorder in Eastern Germany over a time period of eight years. METHOD: A trend analysis was carried out using data from two population surveys conducted in the eastern part of Germany in 1993 and 2001. By means of a fully structured interview psychiatric labelling, causal beliefs, help-seeking and treatment recommendations as well as the desire for social distance was assessed. RESULTS: While there was an increase in the mental health literacy of the public, the desire for social distance from people with major depression and schizophrenia remained unchanged or even increased. CONCLUSIONS: The assumption underlying a number of anti-stigma campaigns, namely that educating people about mental disorders may automatically lead to the improvement of their attitudes towards the mentally ill, appears questionable.

Extreme hyponatremia in an infant with congenital adrenal hypoplasia due to a novel NR0B1 (DAX-1) mutation.

BACKGROUND: Extreme hyponatremia (<105 mmol/l) has rarely been reported in infants. It is potentially life-threatening and requires intensive care treatment. PATIENT: We report on a male infant with absence of weight gain from birth to day 33 of life despite adequate nutrition. On admission serum sodium and potassium were 104 and 5.9 mmol/L respectively. The infant's physical status revealed dehydration, but normal activity with no apparent neurological, circulatory or respiratory impairment. MAIN RESULTS: Global adrenocortical insufficiency was diagnosed and treated with hormonal substitution in addition to intravenous application of fluid, glucose and electrolytes. The rise of serum sodium was carefully monitored and adjusted to a target rate of 0.5 mmol/L/h. X-linked adrenal hypoplasia congenita (X-AHC) was confirmed by the identification of a novel nonsense NR0B1 (DAX-1) mutation (W236X). CONCLUSIONS: X-AHC in infants may present as failure to thrive despite adequate nutrition. Extreme hyponatremia may be associated with little symptoms if developing slowly. Rehydration and slow correction of serum sodium with solutions containing less sodium than normal saline is essential.

[Malassezia furfur colonising the respiratory tract of mechanically ventilated neonates]. / Malassezia furfur im trachealsekret beatmeter frühgeborener.

BACKGROUND: Ventilated neonates are prone to acquire ventilator-associated pneumonia (VAP). Consequently early diagnosis of pneumonia is required. Beside bacteria, fungi are suspected as a cause of VAP. However, fungal colonisation and infection of the lung have not been studied systematically. The purpose of this study was to evaluate pulmonary fungal colonisation in ventilated neonates and premature infants. MATERIALS AND METHODS: 187 tracheal aspirates (TA) from 29 ventilated neonates (23-35 weeks gestational age) were investigated. TAs were evaluated microscopically and by culture. Data were matched with clinical signs of VAP or sepsis. RESULTS: Candida species were not detected in TA or culture. In contrast, Malassezia furfur (Mf), a lipophilic fungus, was detected from the 10th, 21st and 31st postnatal days onwards in TAs of 3 out of 17 extremely prematures (gestational age at birth < 25 weeks). The presence of Mf was associated with clinical deterioration either immediately or a few days after the first positive Mf smear. Topic steroids were more frequently applied to Mf-positive ELBW infants (p = 0.03). In vitro, natural surfactant was demonstrated to be a sufficient substrate for Mf in culture. CONCLUSION AND DISCUSSION: This is the first report on Mf lung colonisation of ELBWI during mechanical ventilation. Because Mf is generally not detected in standard cultures it appears to be an overlooked, potentially pathogenic fungus in prematures. Mf must be considered in the differential diagnosis of VAP in ELBWI exposed to topical steroids, especially when natural surfactant was administered.

Temperature-sensitive formation of chloroplast protrusions and stromules in mesophyll cells of Arabidopsis thaliana.

In leaf mesophyll cells of transgenic Arabidopsis thaliana plants expressing GFP in the chloroplast, stromules (stroma-filled tubules) with a length of up to 20 mum and a diameter of about 400-600 nm are observed in cells with spaces between the chloroplasts. They appear extremely dynamic, occasionally branched or polymorphic. In order to investigate the effect of temperature on chloroplasts, we have constructed a special temperature-controlled chamber for usage with a light microscope (LM-TCC). This LM-TCC enables presetting of the temperature for investigation directly at the microscope stage with an accuracy of +/-0.1 degrees C in a temperature range of 0 degrees C to +60 degrees C. With the LM-TCC a temperature-dependent appearance of chloroplast protrusions has been found. These structures have a considerably smaller length-to-diameter ratio than typical stromules and reach a length of 3-5 mum. At 5-15 degrees C (low temperatures), almost no chloroplast protrusions are observed, but they appear with increasing temperatures. At 35-45 degrees C (high temperatures), numerous chloroplast protrusions with a beaklike appearance extend from a single chloroplast. Interaction of stromules with other organelles has also been investigated by transmission electron microscopy. At 20 degrees C, transverse sections of stromules are frequently observed with a diameter of about 450 nm. A close membrane-to-membrane contact of stromules with the nucleus and mitochondria has been visualised. Golgi stacks and microbodies are found in the spatial vicinity of stromules. At 5 degrees C, virtually no chloroplast protrusions or stromules are observed. At 35 degrees C, chloroplast protrusions are present as broader thylakoid-free stroma-filled areas, resulting in an irregular chloroplast appearance.

Design and construction of a new temperature-controlled chamber for light and confocal microscopy under monitored conditions: biological application for plant samples.

A new light microscope-temperature-controlled chamber (LM-TCC) has been constructed. The special feature of the light microscope-temperature-controlled chamber is the Peltier-element temperature control of a specimen holder for biological samples, with a volume capacity of 1 mL. This system has marked advantages when compared to other approaches for temperature-controlled microscopy. It works in a temperature range of -10 degrees C to +95 degrees C with an accuracy of +/-0.1 degrees C in the stationary phase. The light microscope-temperature-controlled chamber allows rapid temperature shift rates. A maximum heating rate of 12.9 degrees C min(-1) and a maximum cooling rate of 6.0 degrees C min(-1) are achieved with minimized overshoots (