Razor Thin Win: The 1972 Constitution Ballot, Campaign, Ratification & the Courts -- Mae Nan (Robinson) Ellingson & Mick McKeon “In the Crucible of Change”

Beyond the challenge of crafting a new state Constitution that empowered the people and modernized and opened up state and local government in Montana, the Constitutional Convention delegates, as they signed the final document, looked forward to the arduous task of getting it ratified by the electorate in a short ten week period between the end of the convention on March 24 and the ratification election of June 6, 1972. While all 100 delegates signed the draft Constitution, not all supported its adoption. But the planning about how to get it adopted went back to the actions of the Convention itself, which carefully crafted a ballot that kept “hot political issues” from potentially killing the entire document at the polls. As a result, three side issues were presented to the electorate on the ballot. People could vote for or against those side issues and still vote to ratify the entire document. Thus, the questions of legalizing gambling, having a unicameral legislature and retaining the death penalty were placed separately on the ballot (gambling passed, as did the retention of the death penalty, but the concept of a one-house legislature was defeated). Once the ballot structure was set, delegates who supported the new Constitution organized a grassroots, locally focused effort to secure ratification – thought hampered by a MT Supreme Court decision on April 28 that they could not expend $45,000 in public monies that they had set aside for voter education. They cobbled together about $10,000 of private money and did battle with the established political forces, led by the MT Farm Bureau, MT Stockgrowers’ Assn. and MT Contractors Assn., on the question of passage. Narrow passage of the main document led to an issue over certification and a Montana Supreme Court case challenging the ratification vote. After a 3-2 State Supreme Court victory, supporters of the Constitution then had to defend the election results again before the federal courts, also a successful effort. Montana finally had a new progressive State Constitution that empowered the people, but the path to it was not clear and simple and the win was razor thin. The story of that razor thin win is discussed in this chapter by the two youngest delegates to the 1972 Constitutional Convention, Mae Nan Ellingson of Missoula and Mick McKeon, then of Anaconda. Both recognized “Super Lawyers in their later professional practices were also significant players in the Constitutional Convention itself and actively participated in its campaign for ratification. As such, their recollections of the effort provide an insider’s perspective of the struggle to change Montana for the better through the creation and adoption of a new progressive state Constitution “In the Crucible of Change.” Mae Nan (Robinson) Ellingson was born Mae Nan Windham in Mineral Wells, TX and graduated from Mineral Wells High School in 1965 and Weatherford College in Weatherford, TX in 1967. Mae Nan was the youngest delegate at the 1972 Convention from Missoula. She moved to Missoula in 1967 and received her BA in Political Science with Honors from the University of MT in 1970. She was a young widow known by her late husband’s surname of Robinson while attending UM graduate school under the tutelage of noted Professor Ellis Waldron when he persuaded her to run for the Constitutional Convention. Coming in a surprising second in the delegate competition in Missoula County she was named one of the Convention’s “Ten Outstanding Constitutional Convention Delegates,” an impressive feat at such a young age. She was 24 at the time, the youngest person to serve at the ConCon, and one of 19 women out of 100 delegates. In the decade before the Convention, there were never more than three women Legislators in any session, usually one or two. She was a member of the American Association of University Women, a Pi Sigma Alpha political science honorary, and a Phi Alpha Theta historical honorary. At the Convention, she led proposals for the state's bill of rights, particularly related to equal rights for women. For years, Ellingson kept a copy of the preamble to the Constitution hanging in her office; while all the delegates had a chance to vote on the wording, she and delegate Bob Campbell are credited with the language in the preamble. During the convention, she had an opportunity that opened the door to her later career as an attorney. A convention delegate suggested to her that she should go to law school. Several offered to help, but at the time she couldn't go to school. Her mom had died in Texas, and she ended up with a younger brother and sister to raise in Missoula. She got a job teaching, but about a year later, intrigued with the idea of pursuing the law as a career, she called the man back to ask about the offer. Eventually another delegate, Dave Drum of Billings, sponsored her tuition at the UM School of Law. After receiving her JD with Honors (including the Law Review and Moot Court) from the UM Law School Ellingson worked for the Missoula city attorney's office for six years (1977-83), and she took on landmark projects. During her tenure, Missoula became the first city to issue open space bonds, a project that introduced her to Dorsey & Whitney. The city secured its first easement on Mount Sentinel, and it created the trail along the riverfront with a mix of playing fields and natural vegetation. She also helped develop a sign ordinance for the city of Missoula. She ended up working as bond counsel for Dorsey & Whitney, and she opened up the firm's full-fledged Missoula office after commuting a couple of years to its Great Falls office. She was a partner at Dorsey Whitney, working there from 1983 until her retirement in 2012. The area of law she practiced there is a narrow specialty - it requires knowledge of constitutional law, state and local government law, and a slice of federal tax law - but for Ellingson it meant working on great public projects – schools, sewer systems, libraries, swimming pools, ire trucks. At the state level, she helped form the Montana Municipal Insurance Authority, a pooled insurance group for cities. She's shaped MT’s tax increment law, and she was a fixture in the MT Legislature when they were debating equal rights. As a bond lawyer, though, Ellingson considers her most important work for the state to be setting up the Intercap Program that allowed local governments to borrow money from the state at a low interest rate. She has been a frequent speaker at the League of Cities and Towns, the Montana Association of Counties, and the Rural Water Users Association workshops on topics related to municipal finance, as well as workshops sponsored by the DNRC, the Water and Sewer Agencies Coordination Team, and the Montana State University Local Government Center. In 2002, she received an outstanding service award from the Montana Rural Water Users Association. In addition to being considered an expert on Montana state and constitutional law, local government law and local government finance, she is a frequent teacher at the National Association of Bond Lawyers (NABL) Fundamentals of Municipal Bond Law Seminar and the NABL Bond Attorney’s Workshop. For over 30 years Mae Nan has participated in the drafting of legislation in Montana for state and local finance matters. She has served on the Board of Directors of NABL, as Chairman of its Education Committee, was elected as an initial fellow in 1995 to the American College of Bond Counsel, and was recognized as a Super Lawyer in the Rocky Mountain West. Mae Nan was admitted to practice before the MT and US Supreme Courts, was named one of “America’s Leading Business Lawyers” by Chambers USA (Rank 1), a Mountain States Super Lawyer in 2007 and is listed in Best Lawyers in America; she is a member and former Board Member of NABL, a Fellow of the American College of Bond Counsel and a member of the Board of Visitors of the UM Law School. Mae Nan is also a philanthropist who serves on boards and applies her intelligence to many organizations, such as the Missoula Art Museum. [Much of this biography was drawn from a retirement story in the Missoulian and the Dorsey Whitney web site.] Mick McKeon, born in Anaconda in 1946, is a 4th generation Montanan whose family roots in this state go back to the 1870’s. In 1968 he graduated from Notre Dame with a BA in Communications and received a Juris Doctorate degree from the University of Montana Law School in 1971. Right after graduating from law school, Mick was persuaded by his father, longtime State Senator Luke McKeon, and his uncle, Phillips County Attorney Willis McKeon, to run for delegate to Montana’s Constitutional Convention and was elected to represent Deer Lodge, Philipsburg, Powell, and part of Missoula Counties. Along with a coalition of delegates from Butte and Anaconda, he fought through the new Constitution to eliminate the legal strangle hold, often called “the copper collar,” that corporate interests -- the Anaconda Company and its business & political allies -- had over state government for nearly 100 years. The New York Times called Montana’s Constitutional Convention a “prairie revolution.” After helping secure the ratification of the new Constitution, Mick began his practice of law in Anaconda where he engaged in general practice for nearly 20 years. Moving to Butte in 1991, Mick focused has practice in personal injury law, representing victims of negligence and corporate wrongdoing in both Montana district courts and federal court. As such, he participated in some of the largest cases in the history of the state. In 1992 he and his then law partner Rick Anderson obtained a federal court verdict of $11.5 million -- the largest verdict in MT for many years. Mick’s efforts on behalf of injured victims have been recognized by many legal organizations and societies. Recently, Mick was invited to become a member of the International Academy of Trial Lawyers - 600 of the top lawyers in the world. Rated as an American Super Lawyer, he has continuously been named one of the Best Lawyers in America, and an International Assn. of Trial Lawyers top 100 Trial Lawyer. In 2005, he was placed as one of Montana’s top 4 Plaintiff’s lawyers by Law Dragon. Mick is certified as a civil trial specialist by the National Board of Trial Advocacy and has the highest rating possible from Martindale-Hubble. Mick was awarded the Montana Trial Lawyers Public Service Award and provided pro bono assistance to needy clients for his entire career. Mick’s law practice, which he now shares with his son Michael, is limited to representing individuals who have been injured in accidents, concentrating on cases against insurance companies, corporations, medical providers and hospitals. Mick resides in Butte with his wife Carol, a Butte native. Mick, Carol, Michael and another son, Matthew, who graduated from Dartmouth College and was recently admitted to the Montana bar, enjoy as much of their time together in Butte and at their place on Flathead Lake.

Pharmacological inhibition of integrin alphavbeta3 aggravates experimental liver fibrosis and suppresses hepatic angiogenesis

The vitronectin receptor integrin alphavbeta3 promotes angiogenesis by mediating migration and proliferation of endothelial cells, but also drives fibrogenic activation of hepatic stellate cells (HSCs) in vitro. Expecting antifibrotic synergism, we studied the effect of alphavbeta3 inhibition in two in vivo models of liver fibrogenesis. Liver fibrosis was induced in rats by way of bile duct ligation (BDL) for 6 weeks or thioacetamide (TAA) injections for 12 weeks. A specific alphavbeta3 (alphavbeta5) inhibitor (Cilengitide) was given intraperitoneally twice daily at 15 mg/kg during BDL or after TAA administration. Liver collagen was determined as hydroxyproline, and gene expression was quantified by way of quantitative polymerase chain reaction. Liver angiogenesis, macrophage infiltration, and hypoxia were assessed by way of CD31, CD68 and hypoxia-inducible factor-1alpha immunostaining. Cilengitide decreased overall vessel formation. This was significant in portal areas of BDL and septal areas of TAA fibrotic rats and was associated with a significant increase of liver collagen by 31% (BDL) and 27% (TAA), and up-regulation of profibrogenic genes and matrix metalloproteinase-13. Treatment increased gamma glutamyl transpeptidase in both models, while other serum markers remained unchanged. alphavbeta3 inhibition resulted in mild liver hypoxia, as evidenced by up-regulation of hypoxia-inducible genes. Liver infiltration by macrophages/Kupffer cells was not affected, although increases in tumor necrosis factor alpha, interleukin-18, and cyclooxygenase-2 messenger RNA indicated modest macrophage activation. CONCLUSION: Specific inhibition of integrin alphavbeta3 (alphavbeta5) in vivo decreased angiogenesis but worsened biliary (BDL) and septal (TAA) fibrosis, despite its antifibrogenic effect on HSCs in vitro. Angiogenesis inhibitors should be used with caution in patients with hepatic fibrosis.

Lack of galactose-alpha-1,3-galactose expression on porcine endothelial cells prevents complement-induced lysis but not direct xenogeneic NK cytotoxicity

The galactose-alpha-1,3-galactose (alphaGal) carbohydrate epitope is expressed on porcine, but not human cells, and therefore represents a major target for preformed human anti-pig natural Abs (NAb). Based on results from pig-to-primate animal models, NAb binding to porcine endothelial cells will likely induce complement activation, lysis, and hyperacute rejection in pig-to-human xenotransplantation. Human NK cells may also contribute to innate immune responses against xenografts, either by direct recognition of activating molecules on target cells or by FcgammaRIII-mediated xenogeneic Ab-dependent cellular cytotoxicity (ADCC). The present study addressed the question as to whether the lack of alphaGal protects porcine endothelial cells from NAb/complement-induced lysis, direct xenogeneic NK lysis, NAb-dependent ADCC, and adhesion of human NK cells under shear stress. Homologous recombination, panning, and limiting dilution cloning were used to generate an alphaGal-negative porcine endothelial cell line, PED2*3.51. NAb/complement-induced xenogeneic lysis of PED2*3.51 was reduced by an average of 86% compared with the alphaGal-positive phenotype. PED2*3.51 resisted NK cell-mediated ADCC with a reduction of lysis ranging from 30 to 70%. However, direct xenogeneic lysis of PED2*3.51, mediated either by freshly isolated or IL-2-activated human NK cells or the NK cell line NK92, was not reduced. Furthermore, adhesion of IL-2-activated human NK cells did not rely on alphaGal expression. In conclusion, removal of alphaGal leads to a clear reduction in complement-induced lysis and ADCC, but does not resolve adhesion of NK cells and direct anti-porcine NK cytotoxicity, indicating that alphaGal is not a dominant target for direct human NK cytotoxicity against porcine cells.

Transformation of local medicinal knowledge in the Andean Highlands: Case studies from Peru and Bolivia

In the Andean highlands, indigenous environmental knowledge is currently undergoing major changes as a result of various external and internal factors. As in other parts of the world, an overall process of erosion of local knowledge can be observed. In response to this trend, some initiatives that adopt a biocultural approach aim at actively strengthening local identities and revalorizing indigenous environmental knowledge and practices, assuming that such practices can contribute to more sustainable management of biodiversity. However, these initiatives usually lack a sound research basis, as few studies have focused on the dynamics of indigenous environmental knowledge in the Andes and on its links with biodiversity management. Against this background, the general objective of this research project was to contribute to the understanding of the dynamics of indigenous environmental knowledge in the Andean highlands of Peru and Bolivia by investigating how local medicinal knowledge is socially differentiated within rural communities, how it is transformed, and which external and internal factors influence these transformation processes. The project adopted an actor-oriented perspective and emphasized the concept of knowledge dialogue by analyzing the integration of traditional and formal medicinal systems within family therapeutic strategies. It also aimed at grasping some of the links between the dynamics of medicinal knowledge and the types of land use systems and biodiversity management. Research was conducted in two case study areas of the Andes, both Quechua-speaking and situated in comparable agro-ecological production belts - Pitumarca District, Department of Cusco (Southern Peruvian Highlands) and the Tunari National Park, Department of Cochabamba (Bolivian inner-Andean valleys). In each case study area, the land use systems and strategies of 18 families from two rural communities, their environmental knowledge related to medicine and to the local therapeutic flora, and an appreciation of the dynamics of this knowledge were assessed. Data were collected through a combination of disciplinary and participatory action-research methods. It was mostly analyzed using qualitative methods, though some quantitative ethnobotanical methods were also used. In both case studies, traditional medicine still constitutes the preferred option for the families interviewed, independently of their age, education level, economic status, religion, or migration status. Surprisingly and contrary to general assertions among local NGOs and researchers, results show that there is a revival of Andean medicine within the younger generation, who have greater knowledge of medicinal plants than the previous one, value this knowledge as an important element of their way of life and relationship with “Mother Earth” (Pachamama), and, at least in the Bolivian case, prefer to consult the traditional healer rather than go to the health post. Migration to the urban centres and the Amazon lowlands, commonly thought to be an important factor of local medicinal knowledge loss, only affects people’s knowledge in the case of families who migrate over half of the year or permanently. Migration does not influence the knowledge of medicinal plants or the therapeutic strategies of families who migrate temporarily for shorter periods of time. Finally, economic status influences neither the status of people’s medicinal knowledge, nor families’ therapeutic strategies, even though the financial factor is often mentioned by practitioners and local people as the main reason for not using the formal health system. The influence of the formal health system on traditional medicinal knowledge varies in each case study area. In the Bolivian case, where it was only introduced in the 1990s and access to it is still very limited, the main impact was to give local communities access to contraceptive methods and to vaccination. In the Peruvian case, the formal system had a much greater impact on families’ health practices, due to local and national policies that, for instance, practically prohibit some traditional practices such as home birth. But in both cases, biomedicine is not considered capable of responding to cultural illnesses such as “fear” (susto), “bad air” (malviento), or “anger” (colerina). As a consequence, Andean farmers integrate the traditional medicinal system and the formal one within their multiple therapeutic strategies, reflecting an inter-ontological dialogue between different conceptions of health and illness. These findings reflect a more general trend in the Andes, where indigenous communities are currently actively revalorizing their knowledge and taking up traditional practices, thus strengthening their indigenous collective identities in a process of cultural resistance.

What is the Relation between Human Practical Action and an Accompanying Discourse? Discussing the Status of Practical Theory

The article discusses the function of an accompanying discourse in relation to the genesis of human practical action. On the one side, theory cannot be taken as the ground for practical action; practical action is not a realisation of intentions. On the other hand, human practical action is accompanied by series of explanations, justifications, declarations of intent, pre‑ and post-rationalisations, motivations etc. These accompanying discourses seem in one way or the other to be necessary for the actual realisation of human practical action. Following Pierre Bourdieu, it is suggested that an accompanying discourse cannot in a meaningful manner be separated from the human practical action, that practical theory should be regarded not as theory but as part of practice, and that practical theory first of all provides a common language for talking about practice and hence for reproducing a fundamentally arbitrary idea of the genesis of human practical action. Parallels are drawn to the education/formal training of semi-professionals.

Landscapes of Education and the New Role of Social Work International Models of How to Combine Formal, Non-formal and Informal Education

Landscapes of education are a new topic within the debate about adequate and just education and human development for everybody. In particular, children and youths from social classes affected by poverty, a lack of prospects or minimal schooling are a focal group that should be offered new approaches and opportunities of cognitive and social development by way of these landscapes of education. It has become apparent that the traditional school alone does not suffice to meet this need. There is no doubt that competency-based orientation and employability are core areas with the help of which the generation now growing up will manage the start of its professional career. In addition and by no means less important, the development involves individual, social, cultural and societal perspectives that can be combined under the term of human development. In this context, the Capability Approach elaborated by Amartya Sen and Martha Nussbaum has developed a more extensive concept of human development and related it to empirical instruments. Using the analytic concept of individual capabilities and societal opportunities they shaped a socio-political formula that should be adapted in particular to modern social work. Moreover, the Capability Approach offers a critical foil with regard to further development and revision of institutionalised approaches in education and human development.

Limitations: The Centerpiece of Copyright in Distress

After the exclusive rights in copyright have been consolidated in a century-long historical development, limitations and exceptions have become the main instrument to determine the exact scope of copyright. Limitations and exceptions do not merely fine-tune copyright protection. Rather, they balance the interests of authors, rightholders, competitors and end-users in a quadrupolar copyright system. Understanding this is of particular importance in the digital and networked information society, where copyrighted information is not only created and consumed, but constantly extracted, regrouped, repackaged, recombined, abstracted and interpreted. However, serious doubts exist whether the present, historically grown system of limitations adequately balances the interests involved in the information society. Both the closed list of limitations allowed under Art. 5 of the EU Information Society Directive 2001/29/EC and a narrowly interpreted three-step test contained in Arts. 13 TRIPS and 5 (5) of the Information Society Directive appear as obstacles in the way of achieving the appropriate balance needed. This brief article outlines the issues involved which were discussed at the International Conference on “Commons, Users, Service Providers – Internet (Self-) Regulation and Copyright” which took place in Hannover, Germany, on 17/18 March 2010 on the occasion of the launch of JIPITEC.

Gingival recessions and the change of inclination of mandibular incisors during orthodontic treatment

SUMMARY A recent systematic review demonstrated that, overall, orthodontic treatment might result in a small worsening of periodontal status. The aim of this retrospective study was to test the hypothesis that a change of mandibular incisor inclination promotes development of labial gingival recessions. One hundred and seventy-nine subjects who met the following inclusion criteria were selected: age 11-14 years at start of orthodontic treatment (TS), bonded retainer placed immediately after treatment (T₀), dental casts and lateral cephalograms available pre-treatment (TS), post-treatment (T₀), 2 years post-treatment (T₂), and 5 years post-treatment (T₅). Depending on the change of lower incisor inclination during treatment (ΔInc_Incl), the sample was divided into three groups: Retro (N = 34; ΔInc_Incl ≤ -1 degree), Stable (N = 22; ΔInc_Incl > -1 degree and ≤1 degree), and Pro (N = 123; ΔInc_Incl > 1 degree). Clinical crown heights of mandibular incisors and the presence of gingival recessions in this region were assessed on plaster models. Fisher's exact tests, one-way analysis of variance, and regression models were used for analysis of inter-group differences. The mean increase of clinical crown heights (T₀ to T₅) of mandibular incisors ranged from 0.6 to 0.91 mm in the Retro, Stable, and Pro groups, respectively; the difference was not significant (P = 0.534). At T₅, gingival recessions were present in 8.8, 4.5, and 16.3 per cent patients from the Retro, Stable, and Pro groups, respectively. The difference was not significant (P = 0.265). The change of lower incisors inclination during treatment did not affect development of labial gingival recessions in this patient group.

Energy-Efficient Management Of Heterogeneous Wireless Sensor Networks

Various applications for the purposes of event detection, localization, and monitoring can benefit from the use of wireless sensor networks (WSNs). Wireless sensor networks are generally easy to deploy, with flexible topology and can support diversity of tasks thanks to the large variety of sensors that can be attached to the wireless sensor nodes. To guarantee the efficient operation of such a heterogeneous wireless sensor networks during its lifetime an appropriate management is necessary. Typically, there are three management tasks, namely monitoring, (re) configuration, and code updating. On the one hand, status information, such as battery state and node connectivity, of both the wireless sensor network and the sensor nodes has to be monitored. And on the other hand, sensor nodes have to be (re)configured, e.g., setting the sensing interval. Most importantly, new applications have to be deployed as well as bug fixes have to be applied during the network lifetime. All management tasks have to be performed in a reliable, time- and energy-efficient manner. The ability to disseminate data from one sender to multiple receivers in a reliable, time- and energy-efficient manner is critical for the execution of the management tasks, especially for code updating. Using multicast communication in wireless sensor networks is an efficient way to handle such traffic pattern. Due to the nature of code updates a multicast protocol has to support bulky traffic and endto-end reliability. Further, the limited resources of wireless sensor nodes demand an energy-efficient operation of the multicast protocol. Current data dissemination schemes do not fulfil all of the above requirements. In order to close the gap, we designed the Sensor Node Overlay Multicast (SNOMC) protocol such that to support a reliable, time-efficient and energy-efficient dissemination of data from one sender node to multiple receivers. In contrast to other multicast transport protocols, which do not support reliability mechanisms, SNOMC supports end-to-end reliability using a NACK-based reliability mechanism. The mechanism is simple and easy to implement and can significantly reduce the number of transmissions. It is complemented by a data acknowledgement after successful reception of all data fragments by the receiver nodes. In SNOMC three different caching strategies are integrated for an efficient handling of necessary retransmissions, namely, caching on each intermediate node, caching on branching nodes, or caching only on the sender node. Moreover, an option was included to pro-actively request missing fragments. SNOMC was evaluated both in the OMNeT++ simulator and in our in-house real-world testbed and compared to a number of common data dissemination protocols, such as Flooding, MPR, TinyCubus, PSFQ, and both UDP and TCP. The results showed that SNOMC outperforms the selected protocols in terms of transmission time, number of transmitted packets, and energy-consumption. Moreover, we showed that SNOMC performs well with different underlying MAC protocols, which support different levels of reliability and energy-efficiency. Thus, SNOMC can offer a robust, high-performing solution for the efficient distribution of code updates and management information in a wireless sensor network. To address the three management tasks, in this thesis we developed the Management Architecture for Wireless Sensor Networks (MARWIS). MARWIS is specifically designed for the management of heterogeneous wireless sensor networks. A distinguished feature of its design is the use of wireless mesh nodes as backbone, which enables diverse communication platforms and offloading functionality from the sensor nodes to the mesh nodes. This hierarchical architecture allows for efficient operation of the management tasks, due to the organisation of the sensor nodes into small sub-networks each managed by a mesh node. Furthermore, we developed a intuitive -based graphical user interface, which allows non-expert users to easily perform management tasks in the network. In contrast to other management frameworks, such as Mate, MANNA, TinyCubus, or code dissemination protocols, such as Impala, Trickle, and Deluge, MARWIS offers an integrated solution monitoring, configuration and code updating of sensor nodes. Integration of SNOMC into MARWIS further increases performance efficiency of the management tasks. To our knowledge, our approach is the first one, which offers a combination of a management architecture with an efficient overlay multicast transport protocol. This combination of SNOMC and MARWIS supports reliably, time- and energy-efficient operation of a heterogeneous wireless sensor network.

Interferon-α antagonizes IL-3-mediated immunoregulatory functions of human basophils

Human basophils are major inflammatory cells in maintaining chronic allergic asthma. It has been published that interferon-α (IFN-α) improves clinical symptoms of asthma patients. In contrast, IL-3 exacerbates airway inflammation by inducing IL-4, IL-8 and IL-13 secretion from human basophils thus regulating their immunoregulatory functions. Furthermore, IL-3 exceptionally promotes survival of basophils. Here, we assessed cellular response of human basophils treated with IFN-α alone or in combination with IL-3. Our data show that IFN-α enhances apoptosis in purified human blood basophils compared to spontaneous apoptosis of controls or IFN-γ treated cells. Furthermore, we demonstrate that both IFN-α and FasL enhance apoptosis in human basophils with similar efficiency in a rather additive than synergistic way. IFN-α inhibits IL-3-induced survival to a minor degree. Particularly however, it suppresses IL-3-induced de-novo production of IL-8 and IL-13 up to 80%. In contrast, the production of IL-4 is not affected. Analyses of signaling pathways reveal that IFN-α promotes prolonged phosphorylation of STAT1/STAT2. By using a pan-JAK inhibitor the phosphorylation of STAT1/STAT2 is inhibited and most importantly the pro-apoptotic effect of IFN-α is abolished. Although the phosphorylation of p38-MAPK in IFN-α-treated cells is comparable to non-treated cells, inhibition of p-p38 activity abrogates IFN-α-enhanced apoptosis as well. We conclude that IFN-α-enhanced apoptosis is tightly regulated by the cooperation of JAK/STAT and p38-MAPK pathways. Our study identifies IFN-α as a novel inhibitor of IL-3-induced IL-8 and IL-13 production of human basophils. Taken together our study may explain the improved clinical symptoms of asthma patients treated with IFN-α.

Conversion of myoblasts to physiologically active neuronal phenotype.

Repressor element 1 (RE1)-silencing transcription factor (REST)/neuron-restrictive silencer factor (NRSF) can repress several terminal neuronal differentiation genes by binding to a specific DNA sequence (RE1/neuron-restrictive silencer element [NRSE]) present in their regulatory regions. REST-VP16 binds to the same RE1/NRSE, but activates these REST/NRSF target genes. However, it is unclear whether REST-VP16 expression is sufficient to cause formation of functional neurons either from neural stem cells or from heterologous stem cells. Here we show that the expression of REST-VP16 in myoblasts grown under muscle differentiation conditions blocked entry into the muscle differentiation pathway, countered endogenous REST/NRSF-dependent repression, activated the REST/NRSF target genes, and, surprisingly, activated other neuronal differentiation genes and converted the myoblasts to a physiologically active neuronal phenotype. Furthermore, in vitro differentiated neurons produced by REST-VP16-expressing myoblasts, when injected into mouse brain, survived, incorporated into the normal brain, and did not form tumors. This is the first instance in which myoblasts were converted to a neuronal phenotype. Our results suggest that direct activation of REST/NRSF target genes with a single transgene, REST-VP16, is sufficient to activate other terminal neuronal differentiation genes and to override the muscle differentiation pathways, and they suggest that this approach provides an efficient way of triggering neuronal differentiation in myoblasts and possibly other stem cells.

The hylEfm gene in pHylEfm of Enterococcus faecium is not required in pathogenesis of murine peritonitis.

BACKGROUND: Plasmids containing hylEfm (pHylEfm) were previously shown to increase gastrointestinal colonization and lethality of Enterococcus faecium in experimental peritonitis. The hylEfm gene, predicting a glycosyl hydrolase, has been considered as a virulence determinant of hospital-associated E. faecium, although its direct contribution to virulence has not been investigated. Here, we constructed mutants of the hylEfm-region and we evaluated their effect on virulence using a murine peritonitis model. RESULTS: Five mutants of the hylEfm-region of pHylEfmTX16 from the sequenced endocarditis strain (TX16 [DO]) were obtained using an adaptation of the PheS* system and were evaluated in a commensal strain TX1330RF to which pHylEfmTX16 was transferred by mating; these include i) deletion of hylEfm only; ii) deletion of the gene downstream of hylEfm (down) of unknown function; iii) deletion of hylEfm plus down; iv) deletion of hylEfm-down and two adjacent genes; and v) a 7,534 bp deletion including these four genes plus partial deletion of two others, with replacement by cat. The 7,534 bp deletion did not affect virulence of TX16 in peritonitis but, when pHylEfmTX16Δ7,534 was transferred to the TX1330RF background, the transconjugant was affected in in vitro growth versus TX1330RF(pHylEfmTX16) and was attenuated in virulence; however, neither hylEfm nor hylEfm-down restored wild type function. We did not observe any in vivo effect on virulence of the other deletions of the hylEfm-region CONCLUSIONS: The four genes of the hylEfm region (including hylEfm) do not mediate the increased virulence conferred by pHylEfmTX16 in murine peritonitis. The use of the markerless counterselection system PheS* should facilitate the genetic manipulation of E. faecium in the future.

Determining the roles of dendritic cells and ICAM-1 in the transpresentation of IL-15 to CD8 T cells

The maintenance and generation of memory CD8 T cells is dependent on the cytokine IL-15. IL-15 is delivered by a novel mechanism termed transpresentation: IL-15 is presented by a cell expressing IL-15Ralpha to the CD8 T cell which responds via IL-2Rbeta/gammac. The identity of what cells transpresent IL-15 to support the survival and homeostatic proliferation of memory CD8 T cells is unknown. Using a transgenic mouse model that limits IL-15 transpresentation to DCs, I have demonstrated that DCs transpresent IL-15 to CD8 T cells. DCs transpresent IL-15 to CD8 T cells during the contraction of an immune response and also drive homeostatic proliferation of memory CD8 T cells. Additionally, I identified a role for ICAM-1 in promoting homeostatic proliferation. Wt memory CD8 T cells displayed impaired homeostatic proliferation in ICAM-1-/- hosts but not in models of acute IL-15-driven proliferation. In this way, the role of ICAM-1 in IL-15 transpresentation resembles the role for ICAM-1 in antigenpresentation: where antigen or IL-15 is limited, adhesion molecules are important for generating maximal responses. In vitro cultures between CD8 T cells and bone marrowdifferentiated DCs (BMDC) activated with a TLR agonist established a model of proliferation and signaling in CD8 T cells that was dependent on IL-15 transpresentation and required ICAM-1 expression by BMDCs. Regarding the expression of IL-15, I demonstrated that in normal mice it is undetectable without stimulation but is elevated in lymphopenic mice, suggesting a role for T cells in regulating IL-15 expression. Overall, these studies have identified many novel aspects of the interaction between DCs and CD8 T cells that were previously unknown. The study of adhesion molecules in IL-15 transpresentation describes a novel role for these well-known adhesion molecules and it will be interesting for future studies to further characterize this relationship for other IL-15-dependent cell types.

Determining the roles of dendritic cells and ICAM-1 in the transpresentation of IL-15 to CD8 T cells

The maintenance and generation of memory CD8 T cells is dependent on the cytokine IL-15. IL-15 is delivered by a novel mechanism termed transpresentation: IL-15 is presented by a cell expressing IL-15Ralpha to the CD8 T cell which responds via IL-2Rbeta/gammac. The identity of what cells transpresent IL-15 to support the survival and homeostatic proliferation of memory CD8 T cells is unknown. Using a transgenic mouse model that limits IL-15 transpresentation to DCs, I have demonstrated that DCs transpresent IL-15 to CD8 T cells. DCs transpresent IL-15 to CD8 T cells during the contraction of an immune response and also drive homeostatic proliferation of memory CD8 T cells. Additionally, I identified a role for ICAM-1 in promoting homeostatic proliferation. Wt memory CD8 T cells displayed impaired homeostatic proliferation in ICAM-1-/- hosts but not in models of acute IL-15-driven proliferation. In this way, the role of ICAM-1 in IL-15 transpresentation resembles the role for ICAM-1 in antigenpresentation: where antigen or IL-15 is limited, adhesion molecules are important for generating maximal responses. In vitro cultures between CD8 T cells and bone marrowdifferentiated DCs (BMDC) activated with a TLR agonist established a model of proliferation and signaling in CD8 T cells that was dependent on IL-15 transpresentation and required ICAM-1 expression by BMDCs. Regarding the expression of IL-15, I demonstrated that in normal mice it is undetectable without stimulation but is elevated in lymphopenic mice, suggesting a role for T cells in regulating IL-15 expression. Overall, these studies have identified many novel aspects of the interaction between DCs and CD8 T cells that were previously unknown. The study of adhesion molecules in IL-15 transpresentation describes a novel role for these well-known adhesion molecules and it will be interesting for future studies to further characterize this relationship for other IL-15-dependent cell types.