TambiÃ©n fueron publicadas algunas pequeÃ±as citas de autoras como MarÃa Elena Walsh. Por ejemplo, en el primer nÃºmero, se transcribieron unas reflexiones en las que confluye naturalmente la preocupaciÃ³n de la lucha feminista en el marco del contexto polÃtico y cultural de Argentina de los setenta.
Sin embargo, quizÃ¡ el rasgo distintivo y que merece una menciÃ³n destacada (en particular si, como haremos, tomamos luego la lectura que las feministas de los 80 van abk realizar sobre esta experiencia) es la adscripciÃ³n de Persona a los postulados sostenidos por el feminismo radical, tanto por las autoras traducidas y publicadas âla mayorÃa de ellas representantes norteamericanas de dicho feminismoâ, como por las notas elaboradas por militantes argentinas.
El feminismo radical, que, grosso modo, tuvo sus expresiones mÃ¡s destacadas en Estados Unidos, postulÃ³ la existencia de clases sexuales. Este principio le permitiÃ³ identificar con cierta claridad a quiÃ©nes debÃa enfrentar la lucha feminista âconcretamente, al varÃ³n que no estuviera dispuesto a ceder sus beneficios expropiados a las mujeresâ, al tiempo que permitÃa una identificaciÃ³n y unificaciÃ³n de la poblaciÃ³n militante femenina.
En las pÃ¡ginas de Persona, este componente radical se expresÃ³ en frases como esta.
Como seÃ±alÃ¡bamos, el aspecto derivado de este postulado del feminismo radical es una unificaciÃ³n de la poblaciÃ³n femenina, de todas las mujeres hermanadas en la opresiÃ³n por el sexismo y el sistema patriarcal, en definitiva, por los varones, en tanto que beneficiarios de este sistema. Verbigracia, en el segundo nÃºmero de Persona se sostiene que: âFeminismo es la lucha que todo sexo femenino ha emprendido contra la supremacÃa masculina que impide la libre expansiÃ³n de la personalidad femenina, negÃ¡ndole derechos fundamentales y oprimiÃ©ndola con todos los deberesâ (Persona, nÃºm. 2, p. 26; las cursivas son nuestras).
A lo largo de toda la publicaciÃ³n, la caracterizaciÃ³n del sujeto mujer oprimido no admitiÃ³ ninguna otra determinaciÃ³n. Las mujeres compartÃan una realidad que atravesaba a todas las clases y grupos sociales. Afirmaba MarÃa Luisa Bemberg (ufa, en una nota publicada en la revista Claudia en julio de 1973, que:
AsÃ es que Persona sale a la calle âcon el propÃ³sito de informar, analizar y testimoniar sobre la condiciÃ³n de la mujer en nuestra sociedadâ (Persona, nÃºm. 1, p. 3).
Esta comprensiÃ³n e interpelaciÃ³n del sujeto del feminismo en funciÃ³n de su participaciÃ³n en una condiciÃ³n universalizable, la de mujer, resulta graficada en las fotografÃas de las tapas de los dos primeros nÃºmeros de Persona. En el primero se retratÃ³ a 361-302-2570 una muchacha joven, bella y bien arreglada, parada entre una multitud de gente que caminaba las calles cÃ©ntricas de la ciudad, de frente a la cÃ¡mara y con la mirada perdida en el horizonte. En el epÃgrafe al reverso, puede leerse: âEntre la multitud que puebla nuestras calles, se distingue la figura de una nueva mujer. Decidida, estudiosa y trabajadora, ella avanza hacia el porvenir liberada de tabÃºes y prejuicios, y con la seguridad de ser una Personaâ. Algunas investigadoras han seÃ±alado la similitud de esta imagen con cualquier tapa de revistas dirigidas a pÃºblico femenino. En el segundo nÃºmero, en cambio, la tapa es una foto de una mujer adulta, pobre, andina, encorvada, que lleva en sus espaldas a una niÃ±a. El epÃgrafe afirma: âIndÃgena, campesina, mujer y madre, vÃctima de todos los imperialismos, esta boliviana es el sÃmbolo de la mujer latinoamericanaâ. Estas dos tapas son una buena sÃntesis del amplio universo femenino al que Persona buscÃ³ representar de igual manera y, podrÃamos decir, rechazando cualquier diferenciaciÃ³n entre ellas.
De alguna manera, esta bÃºsqueda de dar cuenta de la representaciÃ³n y unificaciÃ³n de todas las integrantes de esta clase sexual terminÃ³ por producir una representaciÃ³n inocente o, si se quiere, liberal, normalizada, urbana y ciudadana de la poblaciÃ³n de mujeres a las que estas feministas procuraban representar. Y es que Persona, que era probable que estuviera cautiva de la realidad cotidiana de sus integrantes y una retÃ³rica de la igualdad al interior del gÃ©nero femenino, concluyÃ³ retratando al centro la situaciÃ³n de las mujeres de clase media y alta que encarnaban las propias militantes feministas, como si fuesen intercambiables con las realidades de otras mujeres. No pudieron, entonces, dar cuenta ni de las mujeres pobres u obreras ni de las jÃ³venes (y no tan jÃ³venes) militantes que se sumaban al proceso de radicalizaciÃ³n polÃtica que vivÃa la Argentina desde mediados de los aÃ±os 60. Al diagonalizar la revista, se hace evidente el riesgo, o la tentaciÃ³n, de la esencializaciÃ³n, un uso de la esencializaciÃ³n que se presenta con la fuerza de las aporÃas que habitan desde sus comienzos el movimiento feminista y la constituciÃ³n de los sujetos que los definen. Por decirlo en tÃ©rminos de Spivak (1989), en muchos pasajes, cuesta discernir la posiciÃ³n enunciativa; es decir, cuando se trata de un esencialismo que propone una representaciÃ³n sustancializada de las mujeres o de la condiciÃ³n femenina, y cuando se estÃ¡ batallando mediante un esencialismo estratÃ©gico que hace un uso polÃtico de la esencialidad.
TambiÃ©n fueron publicadas algunas pequeÃ±as citas de autoras como MarÃa Elena Walsh. Por ejemplo, en el primer nÃºmero, se transcribieron unas reflexiones en las que confluye naturalmente la preocupaciÃ³n de la lucha feminista en el marco del contexto polÃtico y cultural de Argentina de los setenta.
The 270-579-3273 material employed in this study is 6Â mm thick Aluminum alloy 6061-T6. The chemical composition of the base material is given in TableÂ 1. The nano sized reinforcement particles such as Ti2B is used at different volume percentages (vol. %) such as 2, 4 and 8. The average size of the reinforcement particles is 35Â nm and scanning electron microscope (SEM) microstructure of as-received TiB2 nano-particles are shown in Fig.Â 1. The square groove was made tangent to the pin in the advancing side and which is 1Â mm far away from the center line of the tool rotation on the Aluminum alloy 6061-T6 plate. The H13 tool steel having screwed taper pin profile with shoulder diameter of 24Â mm, pin diameter of 8Â mm and 3.5Â mm height was used. The groove opening initially closed by means of the tool which is having shoulder without pin to avoid the escapement of reinforcement particles from groove while processing. Working range of process parameters along the center line used in FSP are presented in TableÂ 2. The FSP is carried out on a Vertical milling machine (Make HMT FM-2, 10 hp, 3000Â rpm) (See Figs.Â 2 and 3).
After FSP, microstructural observations were carried out at the cross section of NZ of surface nano composites normal to the FSP direction, mechanically polished and etched with Keller\’s reagent (2Â ml HF, 3Â ml HCl, 20Â ml HNO3 and 175Â mlÂ H2O) by employing optical microscope (OM). Microhardness tests were carried out at the cross section of NZ of surface hybrid composites normal to the FSP direction, samples with a load of 15Â g and duration of 15Â s using a Vickers digital microhardness tester. The tensile specimens were taken from the surface hybrid composites normal to the FSP direction and made as per ASTM: E8/E8M-011 standard by wire cut Electrical discharge machining to the required dimensions. Wear test is carried out on a pin-on-disk tro-bometer as per ASTM: G99-05 standard. The prismatic pins of 8Â mm dia are cut from the stir zone, where the axis of the pin perpendicular to the FSprocess direction. The EN31 steel having a hardness of 62 HRC is used as a disc. The dia of the sliding track on the disc surface is 100Â mm. The wear test was carrying out under dry-sliding condition with a constant load of 40Â N, disc rotational of speed 650Â rpm and sliding speed of 3.4Â m/s. Wear rate is determined by, Wear rate (mm3/m) is equal to (volume loss/sliding distance).
Results and discussion
The nano composite surface layer by reinforcing TiB2 particles on 6061-T6 Aluminum Alloy via FSP successfully fabricated. Influence of nano-sized reinforcement particles of TiB2 (average size is 35Â nm) on microstructure, mechanical and tribological behaviour of 6061-T6 Al alloy surface nano composite prepared via Friction stir process was studied and the obtained conclusions are:
The author(s) would like to express thanks the authorities of NIT-Warangal, DMRL-Hyderabad and RCIâHyderabad for provided that the facilities to carried-out my research work. One of the authors Dr. A. Devaraju is thankful to the Principal and the management of S R Engineering College, Warangal-Telangana State, for their constant support during this work.
Plastic bonded explosives (PBXs) based on l,3,5,7-tetranitro-1,3,5,7-tetrazocane (HMX) or 2,4,6-triamino-1,3,5- trinitrobenzene (TATB) with various polymer matrices have been formulated in the literature [1â16]. Polymer matrices; Viton A; a vinylidene fluoride – hexafluoropropylene copolymer, Kel-F 800; a vinylidene fluoride – chlorotrifluoroethylene copolymer, polytetrafluoroethylene, Estane 5703; a poly(ester urethane) block copolymer etc. are mainly used for PBXs formulation due to their higher loading density, homogeneity, better dimensional integrity and higher thermal stability than TNT based melt 5187599991 compositions. The role of polymeric matrices is to minimise their sensitivity, improved mechanical and high thermal properties .
Es Ãºtil que en Ã©tica tengamos en cuenta el contexto social mÃ¡s amplio en el que se presentan las situaciones individuales. Si vemos las acciones individuales en el contexto de prÃ¡cticas mÃ¡s amplias, es decir, de modo que no se separen las situaciones y acciones individuales de circunstancias colectivas mÃ¡s amplias, veremos que hay muchos factores que pueden intervenir para que no se respete la decisiÃ³n de una mujer que quiere interrumpir su embarazo âaunque su decisiÃ³n estÃ© basada en que nunca estableciÃ³ una relaciÃ³n afectiva con el embriÃ³n y, en ese sentido, para la Ã©tica del cuidado, no habrÃa nada inmoral en el acto de abortarâ. Sin embargo, visto en un contexto mÃ¡s amplio, la posibilidad de que las cosas vayan mal en las relaciones entre los seres humanos es muy grande: por ejemplo, existe una gran posibilidad de que distintas personas preocupadas por esa situaciÃ³n particular traten de intervenir para impedir lo que ellos ven como una falla en la actitud de cuidado de esa mujer hacia su posible hijo (por ejemplo, por parte de su pareja, de su familia, de los mÃ©dicos que podrÃan ayudarla, etc.). En una sociedad pluralista hay muchas oportunidades de que distintas concepciones del bien, la virtud y el cuidado choquen entre sÃ. No hay garantÃa, 5128726843 partir solo de una Ã©tica del cuidado, de que esto no suceda y de que unas personas traten de intervenir en los asuntos de otras âincluso por razones de preocupaciÃ³n y de cuidadoâ. La Ã©tica del cuidado puede recomendar la tolerancia y el respeto, pero en el mundo real en donde la gente tiene distintas ideas de lo que es correcto o incorrecto moralmente, e incluso distintas concepciones acerca del cuidado, hay muchas formas de que no se respete la decisiÃ³n individual de una mujer de interrumpir su embarazo. Es aquÃ donde se ve la necesidad de algÃºn tipo de garantÃa de fondo por si esto llegara a suceder, constituida por los derechos âque funcionan como una suerte de reclamos universales de protecciÃ³n a ese espacio de decisiÃ³n individualâ. Waldron resalta la importancia de los derechos en esos contextos:
El lenguaje de los derechos al que recurre el feminismo liberal nos ofrece estas estructuras impersonales en las que se pueden basar las relaciones interpersonales de cuidado cuando Ã©stas llegan a fallar o cuando el espacio de la decisiÃ³n individual estÃ¡ en riesgo âcuando estÃ¡, por ejemplo, a merced de los buenos sentimientos e intenciones de otros y de sus diferentes ideas de lo que es lo bueno o incluso el cuidadoâ. El lenguaje de los derechos sirve como una forma de reclamo moral universal para la protecciÃ³n no solo de las relaciones de cuidado, sino tambiÃ©n de garantÃa de fondo en aquellas situaciones en las que las relaciones de cuidado pueden ir mal o simplemente no bastan. Los derechos no reemplazan los lazos afectivos, sino que les pueden servir de precondiciÃ³n, como afirma Waldron, âpor lo menos en un mundo imperfectoâ. AsÃ pues, tal vez el lenguaje de los derechos al que apela el feminismo liberal no dÃ© cuenta de toda la complejidad moral y afectiva en torno a Gyrase una situaciÃ³n de aborto, pero sÃ es una garantÃa para cuando esos lazos no funcionan bien o distintas concepciones del bien y del cuidado llegan a chocar.
Al renunciar a la incorporaciÃ³n de los derechos como un elemento central de la moralidad, la Ã©tica del cuidado radical, por su mismo carÃ¡cter particularista, no tiene modo de hacer un reclamo moral transparticularista en aquellos casos en que no se respete el espacio de decisiÃ³n individual de una mujer que decide realizarse un aborto, sobre todo en aquellos casos en que las distintas concepciones del cuidado choquen. Es por esto que, me parece, el feminismo en general (y no solo el liberal), en su reclamo de respeto a la decisiÃ³n de cada mujer, debe favorecer un enfoque basado en los derechos de las mujeres por sobre uno basado en afectos y relaciones interpersonales de cuidado. Este Ãºltimo no cuenta con el instrumental teÃ³rico para suplir a los derechos morales y las funciones que cumplen. Es por ello que una Ã©tica feminista liberal debe tener primacÃa sobre una Ã©tica del cuidado. Sin embargo, no debe desechar su perspectiva, sino que debe verla como complementaria a su labor, dado que es necesario, para hacer una evaluaciÃ³n moral del aborto âo de lo que sea que estemos evaluando moralmenteâ, no quedarse solo con el discurso de los derechos, porque este puede ser un discurso muy estrecho que borre la complejidad moral de situaciones particulares (cfr. Baier 1994b). Es necesario no perder de vista la riqueza y variedad del vocabulario de afectos, virtudes y cuidado, âcomponentes necesarios de una moralidad adecuadaâ, como afirma Virginia Held. Por eso, la Ã©tica del cuidado no se debe pasar por alto si lo que se quiere es una evaluaciÃ³n completa de la moralidad del aborto â¢
There was an explicit tension at the heart of much of the literature written in the 1980s, which was when MM-102 architecture in Africa and Asia first became the subject of sustained inquiry. On the one hand, these buildings beguiled because they retained an impressive amount of handcrafted detail. That they were also more obviously exotic in both setting and style than metropolitan examples of similar styles only enhanced their appeal. At the height of postmodernism\’s challenge to modernism, it looked as if a return to historicist architecture enriched with ornament was inevitable, and yet there was something slightly dull about simply repeating Georgian certainties Ã la Quinlan Terry. Thus Lutyens\’s work in New Delhi awakened more admiration than did the details of his only slightly more conventionally classical country houses, while his overtly imperial contributions to the center of London were largely ignored (Irving, 1981). Yet the work of Said and Foucault in particular, suggested that the romantic engagement with style suffused with the haze of nostalgia that characterized the first popular surveys of the subject obscured the often very ugly realities of colonialism and its legacy (Morris, 1983). The attention Said and Foucault focused on the relationship between architecture and power made it difficult to continue to hide power relations, especially when they were expressed spatially, behind the discussion of pretty surfaces. Recent work on Europe pointed as well toward the conclusion that architecture was inherently political (Lane, 1968; Vidler, 1990). The earliest scholarship analyzing the relationship between colonial authority and built form, such as Thomas Metcalf\’s An Imperial Vision: Indian Architecture and Britain\’s Raj, focused on stylistic labels and the exterior surfaces of buildings. It made clear that the substitution of the Indo-Saracenic style, in many ways a transposition of the Gothic Revival into Indian conditions, for the classical styles the British had heretofore employed in India was not a sign of respect for indigenous tradition but a shrewd if unsuccessful effort to solidify political power (Metcalf, 1989). More specifically rooted in the particularities of architecture was Mark Crinson\’s perceptive Empire Building: Orientalism and Victorian Architecture (Crinson, 1996). Crinson\’s discussion of the interface between indigenous and imported ways of building set the stage for the emergence of technology transfer as a key theme in the discussion of nonwestern modernism, although much remains to be done (Cody, 2003).
As the enthusiasm for postmodernism gradually faded at the end of the last century, historians turned their attention from the sixteenth through nineteenth century 7277681726 architecture that served as potential sources for new postmodern buildings to the history of nonwestern modernism. What postmodernists had condemned as homogenous postcolonial modernity is now cherished mid-twentieth century modernism that may signal international savoir faire, attentiveness to indigenous precedent, or both. Published in English but often written by natives of the countries under discussion, new studies of this subject have turned the spotlight on the local context of both iconic examples of mid-century modernism and their lesser known, and often far more humble counterparts. In many cases the motive has been to emphasize the modernity of the places that had nurtured modernism in the 1930s and forties when it was under threat in Europe (Nitzan-Shiftan, 2009). The most exciting of these works, however, focused on the indigenous taste for a style that had generally been considered the handiwork of imported European talent. Sibel Bozdogan, for example, demonstrated that an architecture that was often termed the International Style could equally easily serve nationalist goals, as it did in Turkey in the 1930s (Bozdogan, 2001). Meanwhile Crinson showed that, although modern architecture was widely equated with independence, it had also been the architecture of choice for colonial officials in the 1950s (Crinson, 2003). Indeed the same architects, Maxwell Fry and Jane Drew, nearly simultaneously built housing in Chandigarh, the showcase of post-independence India, and in Nigeria, which became independent only in 1960 (Prakash, 2002). Tom Avermaete has further challenged the assumption that modernism was inherently politically and socially progressive. His perceptive study of ATBAT Afrique directly contradicted the rosier interpretation of the International Style in North African advanced by Jean-Louis Cohen and Monique Eleb (Avermaete, 2005; Avermaete et al., 2012; Cohen and Eleb, 2002). Even the recent return of modernism to fashion in the 1990s has received sustained scholarly attention in the case of China (Zhu, 2009).
Molecular taxonomic methods have identified Cryptosporidium hominis (which infects humans) and C. parvum (which infects cattles, humans and other mammals) as the most commonly (573) 328-8498 of Cryptosporidium in surface and wastewater (Paziewska et al., 2007; Smith et al., 2006). An additional Cryptosporidium species of animal origin, C. meleagridis is the third most common species following C. hominis and C. parvum, capable of infecting immuno-compromised humans (Silverlas et al., 2012). Furthermore, C. canis, C. felis, C. suis and C. muris are minor species responsible for human infections (Xiao, 2010) with most cases detected in HIV-positive patients and in children (Cama et al., 2007; Llorente et al., 2007; Thompson et al., 2005; Xiao et al., 2004). C. andersoni has been reported worldwide in post-weened beef and dairy cattle (Olson et al., 2004) and has been implicated as a cause of sporadic human cryptosporidiosis in Australia together with C. fayeri (Waldron et al., 2011).
Giardiasis in humans and most other mammals is caused by Giardia duodenalis. At least eight genotypes of G. duodenalis have been identified (assemblages AâH), however among them, not only assemblages A and B infect humans, but also a wide range of mammalian hosts, making them potential zoonoses. A degree of host-related sub-structuring has been identified within assemblage A, i.e. AI appears to mainly infect animals, AII mainly infects humans, and AIII mainly infects wild ruminants. Assemblages CâH appear generally to be restricted to companion animals, livestock, and rodents while assemblage H so far has only been found in seals and a seagull (Feng and Xiao, 2011).
In a previous study, the prevalence of G. duodenalis in humans and dogs in a rural village in Cambodia was 18.3% (40/218) and 10.6% (10/94) as shown by PCR, respectively. Giardia assemblages AII and BIII of Giardia-positive samples were characterized in humans. G. duodenalis assemblages BIII, C and mix infection between C and D of positive-samples were among the dogs (Inpankaew et al., 2014). In Vietnam, molecular epidemiological studies of the species and genotypes of Giardia and Cryptosporidium infecting humans are few. C. parvum human genotype was found in three HIV patients in Vietnam (Gatei et al., 2003). Mostly non-zoonotic isolates of G. duodenalis (assemblage E) were detected in cattle and pigs at 201 farms located in five provinces around Hanoi in Northern Vietnam (Geurden et al., 2008). Nguyen et al. (2013) found 28/193 pig fecal samples in central Vietnam positive for Cryposporidium oocysts with 12 samples characterized as C.suis and two samples as Cryptosporidium pig genotype II based on 18S rRNA and HSP-70 gene sequence analysis. Fecal samples from cattle in central Vietnam were found positive for C. parvum bovine genotype and C. andersoni (Nguyen et al., 2007). The two non-zoonotic species C. ryanae and C. bovis were detected in native beef calves 2â6months old in Dac Lac province, central Vietnam (Nguyen et al., 2012). Investigating the molecular epidemiology of Giardia and Cryptosporidium in environmental samples can provide important information with regard to the potential sources of infection and likely routes of transmission to humans and animals. Thus, the specific aim of the present study was to evaluate the prevalence and concentrations of Cryptosporidium spp. and Giardia spp. in environmental samples including surface water, compost and fresh vegetables in Hanam, Vienam and to assess potential contamination sources using molecular epidemiological tools. Such knowledge would provide data to aid risk assessment and management measures for preventing contamination of food and water with protozoa in Vietnam.
Materials and methods
The applications of immunofluorescence microscopy on 134 environmental samples revealed 34 samples (25.4%) positive for Giardia spp. and 47 samples for Cryptosporidium spp. (35.0%) (Table 1). In 25 of these samples (18.7%), co-contamination by both pathogens was found.
Proteolytic enzymes and proteases are necessary for the degradation of surrounding proteins and other tissue components and thus play crucial roles in multiple steps of cancer invasion and metastasis (Edwards and Cancer, 1998). Among the proteases, uPAR and cathepsin B are often detected in higher amounts in malignant tumors and have been attributed to contribute major roles in the cancer progression (Alapati et al., 2012; Malla et al., 2012a; Mohamed and Sloane, 2006; Rao, 2003; Smith and Marshall, 2010). Earlier reports indicate that the blockade of uPAR and cathepsin B expression induced a significant reduction in the migration and invasion capabilities of cancer pepstatin (Ahmed et al., 2003; Matarrese et al., 2010; Nalla et al., 2010; Veeravalli et al., 2010; Victor et al., 2011) by effectively abrogating the activation of MAPK signaling (Rabbani et al., 2010; Wegiel et al., 2009; Wu et al., 2008).
In the present study, we studied the effect of shRNA-mediated downregulation of uPAR and cathepsin B (pUC) on 5310 and 4910 non-GICs and GICs either alone or in combination with radiation treatment. Our findings indicate that treating non-GICs and GICs with pUC alone or in combination with radiation reduced the migration of these cells by regulating the JNKâMAPK signaling through the Ras-PI3K pathway in vitro and in vivo. We also observed that a major pool of p-JNK accumulated in the cytoplasm of untreated or irradiated glioma cells while the activated JNK translocated into the nucleus of the non-GICs and GICs treated with pUC alone and in combination with radiation. Further, cytoplasmic p-JNK interacted with adapter proteins of the focal adhesion complex and drove the cells towards an aggressive migratory phenotype.
Materials & methods
Malignant gliomas are extremely lethal and have a 5-year survival rate of less than 3%. Despite aggressive clinical treatment including surgical resection, radiation and chemotherapy, tumor recurrence is essentially universal (Lathia et al., 2011; Stupp et al., 2005). Failure of these regimens might be attributed to the highly infiltrative nature of the glioma cells that reside in the normal 6464537409 at distant locations from the origin of the tumor. Also, the existence of highly resistant, self-replicating glioma-initiating cells (GICs) decreases the success of existing treatment strategies. Approaches that target the invasive capacity of glioma cells as well as the proliferative nature of GICs may significantly improve therapeutic outcomes.
We have previously demonstrated the isolation and characterization of GICs from established cell lines (Alapati et al., 2012; Malla et al., 2012a); we used 5310 and 4910 GICs for the present study. Radiotherapy is a key treatment modality for treating patients with intracranial tumors, but its efficacy is limited by radioresistance and by the promotion of malignant behavior of the cancer cells (Kang et al., 2012; Kil et al., 2012). In our present study, radiation treatment increased the migration of the glioma cells as well as the expression of uPAR and cathepsin B. Elevated levels of uPAR and cathepsin B have been strongly correlated with tumor invasiveness (Besch et al., 2007; Levicar et al., 2003; Rao, 2003; Sevenich et al., 2011), indicating that the cells treated with radiation were adapting towards an aggressive invasive phenotype.
The three core protein kinases of the MAPK family are capable of responding to a number of stimuli to produce specific cellular outcomes. In particular, the precise nature of the extracellular stimuli and the repertoire of molecules available in each cell type can determine the localization, timing, intensity and duration of the activation of each member of the MAPK family (Raman et al., 2007; Turjanski et al., 2007). Several reports indicate the involvement of MAPKs in gene expression, proliferation, motility, metabolism and apoptosis (Cuevas et al., 2007; Dhillon et al., 2007; Huang et al., 2004b; Qi and Elion, 2005). Here, we studied the regulation of glioma cell migration by uPAR and cathepsin B via the MAPK pathway.
The WNT signaling pathway is a primary determinant in assigning an A/P positional identity to NPCs. This instructional cue is imparted early during NPC generation and once this identity is established, it is stable and cannot be altered through exogenous manipulation of the WNT pathway. Using a WNT reporter line, we show that endogenous WNT signaling is highly variable among individual Manumycin A as they acquire a NPC phenotype, with cells of posterior identity expressing WNT reporter activity. In addition to expressing markers of posteriorly fated NPCs, most notably genes of the HOX gene cluster, these cells also express multiple WNT ligands. In contrast, NPCs with anterior identity, as detected by a lack of WNT reporter activity, express multiple WNT antagonists. These differences in expression of WNT agonist and antagonist resemble those observed in the developing neural tube inÂ vivo, with posterior tissues expressing WNT proteins and anterior tissues expressing WNT antagonists such as DKK1 and FRZB (Hashimoto etÂ al., 2000; Leyns etÂ al., 1997).
These opposing WNT signals generate an endogenous gradient of WNT activity, which divides the embryonic neural tube along the A/P axis into distinct progenitor domains, each of which gives rise to specific regionalized neurons (Ciani and Salinas, 2005; Kiecker and Niehrs, 2001; NordstrÃ¶m etÂ al., 2002). These progenitor domains have regionally specific gene-expression profiles and differentiation predispositions despite similar levels of expression of the pan-neural markers SOX1 and SOX2 (Pevny etÂ al., 1998; Wood and Episkopou, 1999; Zappone etÂ al., 2000). Here, we showed that NPCs exhibited a broad range of endogenous WNT activity that conferred specific regionalized fates despite comparable expression levels of SOX1 and SOX2, perhaps mimicking the same developmental events that are seen during early inÂ vivo neural tube development. Therefore, the local WNT microenvironment tightly regulates the WNT activity status and hence the positional identity of NPCs.
A somewhat unexpected implication of these gene-expression patterns is that WNT signaling appears to be acting cell autonomously, with WNT signaling activity restricted to those cells expressing WNT genes. Although WNT signaling activity is present in a graded fashion in these NPC cultures, WNT proteins are acting in an autocrine rather than paracrine manner. Furthermore, expression of WNT antagonists may mute the response in cells near or adjacent to WNT secreting cells. A more careful analysis of this cell-based system will likely yield important mechanistic insights into the dynamic nature of WNT signaling during development.
This restricted WNT signaling activity observed in NPC cultures is consistent with the notion that WNT proteins act locally (Habib etÂ al., 2013) and exhibit minimal, if any, extracellular diffusion. A recent study demonstrated that flies expressing an engineered membrane-tethered Wingless (a fly WNT protein) are viable and normally patterned, suggesting that the spread of Wingless is dispensable for patterning and growth (Alexandre etÂ al., 2014). Similarly, in our cell-based system, WNT proteins act locally and do not signal to distant cells. In addition, expression of WNT antagonists in the WNTâ populations may act to block paracrine WNT signaling activity. This local WNT activity is not the result of the physical separation of distinct WNT expressing domains, since this localized activity is retained in a mixed and seemingly homogeneous cell culture system.
While endogenous WNT signaling activity is a major source of heterogeneity among individual NPCs, exogenous manipulation of this signaling pathway can be exploited to impart specific positional identities to NPCs during their generation from hPSCs, thereby reducing cellular heterogeneity. Activation of WNT signaling withÂ purified WNT3a protein or a GSK-3Î² inhibitor (CHIR98014) led to the generation of NPCs with a hindbrain/spinal cord identity, whereas inhibition of WNT signaling with a PORCN inhibitor (IWP2) to block endogenous WNT protein processing led to the generation of NPCs with a forebrain phenotype. As shown in this and other studies (Li etÂ al., 2009; Pankratz etÂ al., 2007), in the absence of any WNT pathway manipulations, NPCs generated form hPSCs are generally biased toward an anterior fate, suggesting that endogenous WNT signaling in these culture systems is relatively low and insufficient to promote posterior fates. Consequently, ectopic activation of WNT signaling produces a prominent shift from an anterior to a posterior fate. In contrast, in the absence of WNT signaling (through IWP2 addition), the relative increase of anterior-related markers, though statistically significant, is less pronounced.
The mechanisms underlying the functional improvement after NSPC transplantation remain unexplained, which has hampered the establishment of therapeutic protocols for the treatment of SCI (Lindvall and Kokaia, 2010). Several possible explanations for the efficacy of the engrafted NSPCs have been suggested, such as neural cell replacement, remyelination, growth support, neuroprotection, and immunomodulation (Volarevic etÂ al., 2013). However, little is known about the therapeutic effects ofÂ the engrafted NSPCs on the disrupted local neural networks in the injured spinal cord. In this study, we focused on the propriospinal system, the intraspinal neural networks connecting each spinal cord segment (Flynn etÂ al., 2011). This system has received a lot of attention as being important for spontaneous functional recovery after incomplete SCI. For example, Courtine and colleagues demonstrated that a pronounced functional recovery occurred following the spontaneous reorganization of propriospinal circuits without the restoration of the direct projections from the 2098564202 to the spinal motor neurons in a rat model of incomplete SCI. Because the extent of spontaneous recovery after SCI is associated with the amount of spared propriospinal circuits, little functional recovery was observed after severe SCI, in which there were few spared host neurons and spared propriospinal circuits (Figures 1 and 2; Conta and Stelzner, 2004; Courtine etÂ al., 2008). In this paper, we showed that the reorganization of propriospinal circuits was promoted by the engrafted NSPCs through synapse formation between the engrafted NSPCs and the host neurons (Figures 4, 5, and 6). In addition, selective ablation of the host neurons abolished theÂ synaptogenic potential of the engrafted NSPCs and negated the efficacy of the reorganization of propriospinal circuits. These results suggest the importance of functional improvement due to the accelerated reorganization of propriospinal circuits by NSPC transplantation after SCI. Indeed, the most prominent effect of NSPC transplantation was observed within 2Â weeks after transplantation, and this phase was consistent with the spontaneous recovery process in the control groups (FigureÂ 1).
To date, fluorescence-activated cell sorting (FACS) or translating ribosomal affinity purification (TRAP) techniques mainly have been used for cell-specific profiling of engrafted cells (Doyle etÂ al., 2008; Lobo etÂ al., 2006). Although these techniques have broad utility, the LMD procedure provided several advantages over these methods in this study. First, LMD makes it possible to capture neuronal cells even in the adult CNS. FACS and TRAP are not recommended for the analysis of mature neural tissues, because myelin and tight extracellular matrix proteins interfere with the isolation of neurons. To apply FACS and TRAP to isolate neuronal cells from the adult CNS, enzymatic dissociation is required to eliminate these interfering substances; however, this enzymatic method often results in a loss of cells, including their mRNAs and proteins (Garg etÂ al., 2014). Second, LMD enables the capture of neuronal cells while maintaining their distinctive structures, such as long axons and dendrites. Many synaptic mRNAs are localized throughout the axonal and dendritic terminals, which are hundreds of micrometers distant from the neuronal cell body. For instance, representative postsynaptic mRNAs, such as Psd95 and Homer1, are distributed widely from the neuronal cell body to the dendritic terminals (Cajigas etÂ al., 2012). Additionally, presynaptic mRNAs, such as Synaptophysin and Synaptotagmin, are present exclusively in the developing growth cone, a specialized structure at the tip of the extending axon (Zivraj etÂ al., 2010). Although the LMD method used in this study did not have sufficient precision in terms of the ability to dissect the axons and dendrites, we successfully identified the gene expression of these synaptic mRNAs in the engrafted NSPCs. More precise dissection with LMD must be developed for investigating distinct distribution of synaptic mRNAs in neuronal cells.
The capacity to delay cell-cycle progression at the G1/S transition is central to tumor suppression by RB proteins, predominantly via interaction with, and inhibition of, the E2F family of S-phase transcriptional activators. In Drosophila, the role of Rbf proteins in cell-cycle regulation is considerably less complex than for mammals, with just two E2F subunits (compared with at least eight in mammals) and one DP cofactor (compared with two in mammals) (Dynlacht etÂ al., 1994; van den Heuvel and Dyson, 2008). Rbf and RB1 share capacity to bind to E2F transcriptional activators, similarly RBL1/p107, RBL2/p130, and Rbf2 bind E2F repressor complexes (Du and Pogoriler, 2006). Drosophila E2F1 activates transcription by forming heterodimers with the DP transcriptional cofactor. In theÂ absence of developmental growth signals, hypophosphorylated Rbf represses E2F-mediated transcription byÂ binding and blocking the transcriptional activation domain of E2F/DP (Giacinti and Giordano, 2006). In response to mitogenic signals, G1-S Cyclin/cyclin-dependent kinase (CDKs) (e.g., CycD and CycE) can hyperphosphorylate Rbf, releasing the E2F1-DP complex to promote S-phase gene transcription (reviewed in Giacinti and Giordano, 2006). Flies have just one CDK inhibitor, Dacapo (Dap), which selectively inhibits CycE/Cdk2, but not CycD/Cdk4 (de Nooij etÂ al., 1996).
The Drosophila testis provides a system for analysis of gene function in two distinct cell populations derived from adjacent stem cell types (the germline and somatic lineage) within their endogenous niche. The testis produces sperm throughout the lifetime of the adult male fly. From the L1 stage, the stem cell niche is composed of a cluster of somatic (205) 897-2573 (the hub) that supports two stem cell populations: the germline stem cells (GSCs) and the somatic stem cells, also known as cyst stem cells (CySCs) (GÃ¶nczy and DiNardo, 1996; Hardy etÂ al., 1979). Each GSC is enclosed by two CySCs, and both populations undergo asymmetric divisions to (1) maintain the stem cell pool and (2) differentiate into gonialblast daughter or somatic cyst cells, respectively (Fuller and Spradling, 2007; Hardy etÂ al., 1979; Yamashita etÂ al., 2003) (Figures 1A and 1B). The gonialblast exits the niche enclosed by a pair of cyst cells and, after four rounds of transit-amplifying (TA) mitotic divisions with incomplete cytokinesis, generates a 16-cell spermatogonial cyst (Hardy etÂ al., 1979). Upon further growth and differentiation, spermatogonial cysts develop into spermatocytes, which undergo meiosis to produce sperm (Fuller and Spradling, 2007) (Figures 1A and 1B). Here, we demonstrate that although Rbf mutants display cell-cycle exit and differentiation defects in both the germline and somatic lineages, Rbf function was only required in the somatic lineage for testes development. Thus, Rbf function in the somatic cell lineage is required non-autonomously for regulating cell-cycle exit and differentiation in the germline.
Analysis of EMS-induced male-sterile mutant collections suggest most alleles elicit meiosis or spermiogenesis phenotypes, but relatively few hits disrupt the stem cell niche (Wakimoto etÂ al., 2004). This is not surprising, as many stem cell determinants and signaling pathways essential to niche function also have critical functions in earlier development, hence loss-of-function alleles are associated with embryonic or larval lethality and will be absent from fertility screens. In contrast, the unique features of meiotic cells (only spermatocytes and oocytes undergo meiosis) or post-meiotic spermatids often derive from factors specific to these processes. Thus, although several genetic screens for male-sterile alleles have identified genes that function in the stem cell niche to regulate maintenance, proliferation, and differentiation of GSCs and/or somatic stem cells, these are often hypomorphic alleles (Castrillon etÂ al., 1993; Kiger etÂ al., 2001).
The first prospective study of spinal cord injury (SCI) prevalence in the USA revised the estimated number of individuals living with SCI upward by 5-fold to 1.3 million (Christopher and Dana Reeve Foundation, 2008). The average age at time of SCI is 34 years, resulting in a lifetime of paralysis associated with a host of medical complications. The impact of SCI in economic terms is highly disproportionate to the incidence of injury, rising to an average lifetime cost of several million dollars for individuals sustaining high-level cervical injuries. Critically, the majority of clinical SCI cases are at the cervical level (52.4%) (Christopher and Dana Reeve Foundation, 2008), making potential therapeutic interventions in cervical SCI rodent models a high priority.
Selection of a target clinical p-Cresyl sulfate is one key to translation of cell therapeutics. Two critical variables are treatment timing and vertebral level (thoracic versus cervical), both of which affect the incidence of spontaneous recovery in man (Fawcett etÂ al., 2007). Rodent contusion models reproduce the principal pathophysiological features of clinical SCI with sensitive and relevant outcome measures (Stokes and Jakeman, 2002; Nishi etÂ al., 2007). With respect to timing, the timeline of pathophysiological events following SCI in animal models versus the human condition is debatable; although many suggest that transplantation 9Â days post-injury (DPI) in the rodent corresponds to the sub-acute clinical setting, while transplantation 30â60 DPI corresponds to the early chronic clinical setting (Houle and Tessler, 2003; Fawcett etÂ al., 2007). Focusing enrollment for an SCI trial on chronic (>3Â months post-SCI) cervical SCI subjects, compared with acute thoracic subjects, could reduce the enrollment required to attain statistical power to discriminate an improvement of 10 AIS (American Spinal Injury Association Impairment Scale) motor points dramatically from 250 to 25 AIS A subjects or from 1,100 to 50 AIS B subjects (Fawcett etÂ al., 2007). Further, the larger pool of chronic SCI individuals may facilitate subject accrual, while an increased delay between injury and enrollment may improve the informed consent process (Anderson and Cummings, 2016).
With respect to vertebral level, there are compelling reasons to drive toward clinical trials focused on cervical SCI inÂ more chronic cases. Many, however, have cautioned against proceeding to clinical trial for cervical SCI based on preclinical data in thoracic SCI models (Kwon etÂ al., 2013). One reason for hesitation is increased recognition that cervical and thoracic injuries have a number of profound differences. For example, functional motor impairment across levels changes due to the anatomical characteristics of the spinal cord. Disruption of spinal circuitry due to systemic autonomic and immune effects is also level specific. Autonomic dysreflexias, particularly abnormal cardiovascular control, affect 50%â70% of human SCI patients with injury above T6 (Krassioukov and Claydon, 2006) but are rare when the injury is below transforming factor level. Accordingly, the impact of modulation of sprouting or connectivity via cell transplantation therapies could exert unanticipated effects in the case of high thoracic and cervical SCI, which would not be evident in low thoracic SCI models. In parallel, disruption of descending sympathetic outflow specifically associated with cervical and high-level SCI has been shown to exert clinically significant and chronic splenic atrophy and immune suppression (Lucin etÂ al., 2007; Zhang etÂ al., 2013). Of note, SCI subjects in most cell therapeutic clinical trials would receive at least transient pharmacological immunosuppressive agents. In summary, these data demonstrate that injury level is a key variable in establishing not only efficacy but also safety in preclinical testing of investigational agents.