bluuebunny/crossref_metadata_2025 huggingface dataset:
Differentiation of Odontoblast-Like Cells From Mouse Induced Pluripotent Stem Cells by Pax9 and Bmp4 Transfection
Authors: Seki Daisuke , Takeshita Nobuo , Oyanagi Toshihito , Sasaki Shutaro , Takano Ikuko , Hasegawa Masakazu , Takano-Yamamoto Teruko
Published: July 2015

The field of tooth regeneration has progressed in recent years, and human tooth regeneration could become viable in the future. Because induced pluripotent stem (iPS) cells can differentiate into odontogenic cells given appropriate conditions, iPS cells are a potential cell source for tooth regeneration. However, a definitive method to induce iPS cell-derived odontogenic cells has not been established. We describe a novel method of odontoblast differentiation from iPS cells using gene transfection. We generated mouse iPS cell-derived neural crest-like cells (iNCLCs), which exhibited neural crest markers. Next, we differentiated iNCLCs into odontoblast-like cells by transfection of Pax9 and Bmp4 expression plasmids. Exogenous Pax9 upregulated expression of Msx1 and dentin matrix protein 1 (Dmp1) in iNCLCs but not bone morphogenetic protein 4 (Bmp4) or dentin sialophosphoprotein (Dspp). Exogenous Bmp4 upregulated expression of Msx1, Dmp1, and Dspp in iNCLCs, but not Pax9. Moreover, cotransfection of Pax9 and Bmp4 plasmids in iNCLCs revealed a higher expression of Pax9 than when Pax9 plasmid was used alone. In contrast, exogenous Pax9 downregulated Bmp4 overexpression. Cotransfection of Pax9 and Bmp4 synergistically upregulated Dmp1 expression; however, Pax9 overexpression downregulated exogenous Bmp4-induced Dspp expression. Together, these findings suggest that an interaction between exogenous Pax9- and Bmp4-induced signaling modulated Dmp1 and Dspp expression. In conclusion, transfection of Pax9 and Bmp4 expression plasmids in iNCLCs induced gene expression associated with odontoblast differentiation, suggesting that iNCLCs differentiated into odontoblast-like cells. The iPS cell-derived odontoblast-like cells could be a useful cell source for tooth regeneration.

Significance

It has been reported that induced pluripotent stem (iPS) cells differentiate into odontogenic cells by administration of recombinant growth factors and coculture with odontogenic cells. Therefore, they can be potential cell sources for tooth regeneration. However, these previous methods still have problems, such as usage of other cell types, heterogeneity of differentiated cells, and tumorigenicity. In the present study, a novel method to differentiate iPS cells into odontoblast-like cells without tumorigenicity using gene transfection was established. It is an important advance in the establishment of efficient methods to generate homogeneous functional odontogenic cells derived from iPS cells.
Drug Transporter-Mediated Protection of Cancer Stem Cells From Ionophore Antibiotics
Authors: Boesch Maximilian , Zeimet Alain G. , Rumpold Holger , Gastl Guenther , Sopper Sieghart , Wolf Dominik
Published: July 2015

Ionophore antibiotics were reported to selectively kill cancer stem cells and to overcome multidrug resistance, but mechanistic studies of the significance of drug transporters for treatment with these compounds are lacking. We applied chemosensitivity testing of well-characterized human cancer cell lines to elaborate on whether drug transporters are involved in protection from the cytotoxic effects of the ionophore antibiotics salinomycin and nigericin. Our experiments demonstrated that ionophore antibiotics were ineffective against both stem-like ovarian cancer side population cells (expressing either ABCB1 or ABCG2) and K562/Dox-H1 cells, which constitute a genetically defined model system for ABCB1 expression. Considering that cancer stem cells often express high levels of drug transporters, we deduced from our results that ionophore antibiotics are less suited to cancer stem cell-targeted treatment than previously thought.

Significance

Ionophore antibiotics such as salinomycin have repeatedly been shown to target cancer stem and progenitor cells from various tumor entities. Meanwhile, cancer stem cell (CSC)-selective toxicity of ionophore antibiotics seems to be a commonly accepted concept that is about to encourage their clinical testing. This study provides data that challenge the concept of targeted elimination of CSC by ionophore antibiotics. Stem-like ovarian cancer side population (SP) cells expressing high levels of ABC drug transporters are shown to largely resist the cytotoxic effects of salinomycin and nigericin. Furthermore, using a small interfering RNA-based knockdown model specific for ABCB1, this study demonstrates that ABC drug transporters are indeed causally involved in mediating protection from ionophore antibiotics. Considering that it is a hallmark of CSCs to exhibit drug resistance conferred by ABC drug transporters, it must be deduced from these results that CSCs may also be protected from ionophore antibiotics by means of drug-transporter mediated efflux.
Perfil dos pacientes submetidos à colonoscopia em um hospital universitário
Authors: Borges Marcacini Stephania Ferreira , Barichello Elizabeth , Barbosa Maria Helena
Published: July 2015

Esta investigación tuvo como objetivo analizar las características sociodemográficas y clínicas, tabaquismo, alcoholismo, práctica de ejercicios físicos de pacientes sometidos a colonoscopia, en el período de febrero a junio de 2012. Se ha observado que en los 84 pacientes entrevistados los síntomas de mayor prevalencia que llevaron a la indicación de la Colonoscopia fueron el dolor abdominal con 39 (46,4%). De los entrevistados cinco (6%) presentaron cáncer colorrectal (CCR), y la historia familiar de esta neoplasia fue relatada por 18 (21,4%) personas y la neoplasia en cuestión apareció en cinco (6%) de los encuestados, el tabaquismo estuvo presente en 17 (20,2%) de los pacientes y el consumo de bebida alcohólica fue negada por 26 (31%) de los entrevistados. En relación a la práctica de actividad física 71 (84,5%) de los pacientes eran insuficientemente activos.
When was the Sun Pyramid Built? Maintaining the Status Quo at Teotihuacan, Mexico
Authors: Rebecca Sload
Published: July 2015

The traditional view based on ceramics is that construction of the Sun Pyramid at Teotihuacan commenced in the first century A .D. Likewise, radiocarbon dates from the artificial cave beneath the Pyramid indicate that it was created at about the same lime. Both Pyramid and cave are seen as having a role in the founding of the city. Recent excavation inside the Pyramid produced radiocarbon dates that cluster in the mid-third century A.D. Members of the Sun Pyramid Project interpreted the dates as representing initial construction of the structure, moving it significantly later in time than previously thought. They also reinterpreted the dates for the construction of the cave, making it contemporaneous with the revised Pyramid construction. This paper adds radiocarbon dates from the cave to the original set and employs Bayesian analysis. The initial interpretation is supported: the dates reflect a cycle of cave creation through termination that began in the midfirst century and lasted about 200 years. I interpret the dates from the Pyramid as reflecting ritual associated with cave termination and a concomitant redefinition of the Pyramid that involved architectural modifications and tunneling. Pyramid and cave dates are reconciled with each other, with ceramics, and with the ceramic chronology. The traditional timing of first century Pyramid construction is maintained, along with its social, political, and economic implications.
Pottery Vessel Function and Foodways at El Chorro de Maíta, Cuba
Authors: Knight Vernon James , Rojas Roberto Valcarcel
Published: July 2015

Our approach assumes that pots are tools, containers whose performance characteristics are adjusted to their primary uses. Traditional agricultural peoples generally distinguish among multiple vessel shapes that have distinct intended uses. In this article, we present afunctional analysis of vessel shapes and sizes performed on a sample of 160 rim sherds from the site of El Chorro de Malta, Cuba. These were assigned to 13 defined vessel shapes, most of which displayed more than one size mode based on estimated orifice diameters. The majority of specimens from El Chorro de Malta are low-profile, composite-contour bowls made in medium and large size modes, followed in frequency by simple-contour bowls and plates. Late Ceramic Age peoples in the Greater Antilles are historically documented as participating in the manioc breadcake-cassareep- stewpot foodway common to much of the tropical lowlands of northern South America. Consequently, it should be possible to show how the container assemblage of El Chorro de Malta is adapted to the requirements of that foodway. We note that, ethnographically, the elaborate processing of bitter manioc itself to produce breadcakes as a staple food does not necessarily require pottery vessels at all. Nonetheless, common stewing as a key component of the foodway—including the production of the condiment called cassareep in the Guianas—can require several containers with potentially distinct performance requirements: one to collect the juice below the sleeve press, another to reduce the expressed juice to the thickened sauce over afire, and a third, the stewpot itself, adapted to simmering vegetable and meat stews. We provisionally suggest that some of the most common shape-size classes at El Chorro de Malta are suited to producing and serving stews and cassareep, the traditional complement to eating manioc breadcake.
Angry Old Women: Peggy Mount and the Performance of Female Ageing in the British Sitcom
Authors: Mortimer Claire
Published: July 2015

This article examines the mature woman as sitcom harridan, exploring Peggy Mount's performance as Cockney matriarch Ada Larkin in The Larkins (1958–60, 1963–64). In terms of female identity, the menopause can be represented as the onset of decline and invisibility, with characters such as the mother-in-law and nag used to ridicule the mature woman, rendered grotesque and abject. The harridan is a problematic figure enjoying licence and authority, whilst articulating issues regarding gender, class and age. Mount's performance style is animated by excessive verbosity and physicality, rooted in working-class culture and denned by unruliness.
An Extra Slice of Jo Brand
Authors: Lockyer Sharon
Published: July 2015

In this candid interview Jo Brand, one of Britain's most popular and successful comedians, discusses her comedy career. Brand shares experiences and critical reflections on a wide range of comedy, from her first stand-up gig in the 1980s through to presenting the Great British Bake Off: An Extra Slice (2014–present). She discusses the inspiration and motivation for her comic material, explains the highs and lows of working in television comedy and offers pragmatic advice for comedians wishing to pursue a career in comedy today.
Using proteomics to probe neurons
Authors: Kim Yunee , Kislinger Thomas
Published: July 2015

Advances in mass spectrometry-based proteomics have allowed researchers to quantify the abundances of the different forms of three closely related proteins in the neurons of mice.
Shock wave synthesis of amino acids from solutions of ammonium formate and ammonium bicarbonate
Authors: Suzuki Chizuka , Furukawa Yoshihiro , Kobayashi Takamichi , Sekine Toshimori , Nakazawa Hiromoto , Kakegawa Takeshi
Published: July 2015

The emergence of life's building blocks, such as amino acids and nucleobases, on the prebiotic Earth was a critical step for the beginning of life. Reduced species with low mass, such as ammonia, amines, or carboxylic acids, are potential precursors for these building blocks of life. These precursors may have been provided to the prebiotic ocean by carbonaceous chondrites and chemical reactions related to meteorite impacts on the early Earth. The impact of extraterrestrial objects on Earth occurred more frequently during this period than at present. Such impacts generated shock waves in the ocean, which have the potential to progress chemical reactions to form the building blocks of life from reduced species. To simulate shock‐induced reactions in the prebiotic ocean, we conducted shock‐recovery experiments on ammonium bicarbonate solution and ammonium formate solution at impact velocities ranging from 0.51 to 0.92 km/s. In the products from the ammonium formate solution, several amino acids (glycine, alanine, ß‐alanine, and sarcosine) and aliphatic amines (methylamine, ethylamine, propylamine, and butylamine) were detected, although yields were less than 0.1 mol % of the formic acid reactant. From the ammonium bicarbonate solution, smaller amounts of glycine, methylamine, ethylamine, and propylamine were formed. The impact velocities used in this study represent minimum cases because natural meteorite impacts typically have higher velocities and longer durations. Our results therefore suggest that shock waves could have been involved in forming life's building blocks in the ocean of prebiotic Earth, and potentially in aquifers of other planets, satellites, and asteroids.
Effects of inherited cores and magmatic overgrowths on zircon (U‐Th)/He ages and age‐eU trends from Greater Himalayan sequence rocks, Mount Everest region, Tibet
Authors: Orme Devon A. , Reiners Peter W. , Hourigan Jeremy K. , Carrapa Barbara
Published: July 2015

Previous constraints on the timing and rate of exhumation of the footwall of the South Tibetan detachment system (STDS) north of Mount Everest suggest rapid Miocene cooling from ∼ 700°C to 120°C between ∼14–17 Ma. However, 25 new single grain zircon He ages from leucogranites intruding Greater Himalayan Sequence rocks in the footwall of the STDS are between 9.9 and 15 Ma, with weighted mean ages between 10 and 12 Ma. Zircon grains exhibit a positive correlation between age and effective uranium (eU). Laser ablation zircon U‐Pb geochronology, detailed SEM observations, and laser ablation depth‐profiling of these zircons reveal low‐eU 0.5–2.5 Ga inherited cores overgrown by high‐eU 17–22 Ma rims. This intragranular zonation produces ages as much as 32% too young when a standard alpha‐ejection correction assuming uniform eU distribution is applied. Modeling of the effects of varying rim thickness and rim eU concentration on the bulk grain eU and alpha‐ejection correction suggests that zonation also exerts the primary control on the form of the age‐eU correlation observed. Application of grain‐specific zonation‐dependent age corrections to our data yields zircon He ages between 14 and 17 Ma, in agreement with AFT and 40Ar/39Ar ages. Growth of magmatic rims followed by cooling to < 120°C within 1–6 million years supports rapid tectonic exhumation associated with slip along the STDS in the Miocene. This study highlights the importance of characterizing parent nuclide zonation in zircon He studies which seek to understand the timing of exhumation along exhumed crustal blocks.
Anomalously strong observations of PKiKP/PcP amplitude ratios on a global scale
Authors: Waszek Lauren , Deuss Arwen
Published: July 2015

The inner core boundary marks the phase transition between the solid inner core and the fluid outer core. As the site of inner core solidification, the boundary provides insight into the processes generating the seismic structures of the inner core. In particular, it may hold the key to understanding the previously observed hemispherical asymmetry in inner core seismic velocity, anisotropy, and attenuation. Here we use a large PKiKP‐PcP amplitude ratio and travel time residual data set to investigate velocity and density contrast properties near the inner core boundary. Although hemispherical structure at the boundary has been proposed by previous inner core studies, we find no evidence for hemispheres in the amplitude ratios or travel time residuals. In addition, we find that the amplitude ratios are much larger than can be explained by variations in density contrast at the inner core boundary or core‐mantle boundary. This indicates that PKiKP is primarily observed when it is anomalously large, due to focusing along its raypath. Using data in which PKiKP is not detected above the noise level, we calculate an upper estimate for the inner core boundary (ICB) density contrast of 1.2 kg m−3. The travel time residuals display large regional variations, which differ on long and short length scales. These regions may be explained by large‐scale velocity variations in the F layer just above the inner core boundary, and/or small‐scale topography of varying magnitude on the ICB, which also causes the large amplitudes. Such differences could arise from localized freezing and melting of the inner core.
Investigating flow properties of partially cemented fractures in Travis Peak Formation using image‐based pore‐scale modeling
Authors: Tokan‐Lawal Adenike , Prodanović Maša , Eichhubl Peter
Published: July 2015

Natural fractures can provide preferred flow pathways in otherwise low‐permeability reservoirs. In deep subsurface reservoirs including tight oil and gas reservoirs, as well as in hydrothermal systems, fractures are frequently lined or completely filled with mineral cement that reduces or occludes fracture porosity and permeability. Fracture cement linings potentially reduce flow connectivity between the fracture and host rock and increase fracture wall roughness, which constricts flow. We combined image‐based fracture space characterization, mercury injection capillary pressure and permeability experiments, and numerical simulations to evaluate the influence of fracture‐lining cement on single‐phase and multiphase flows along a natural fracture from the Travis Peak Formation, a tight gas reservoir sandstone in East Texas. Using X‐ray computed microtomographic image analysis, we characterized fracture geometry and the connectivity and geometric tortuosity of the fracture pore space. Combining level set method‐based progressive quasistatic and lattice Boltzmann simulations, we assessed the capillary‐dominated displacement properties and the (relative) permeability of a cement‐lined fracture. Published empirical correlations between aperture and permeability for barren fractures provide permeability estimates that vary among each other, and differ from our results, vary by several orders of magnitude. Compared to barren fractures, cement increases the geometric tortuosity, aperture variation of the pore space, and capillary pressure while reducing the single‐phase permeability by up to 2 orders of magnitude. For multiphase displacement, relative permeability and fluid entrapment geometry resemble those of porous media and differ from those characteristic of barren fractures.
Seasonal advection of Pacific Equatorial Water alters oxygen and pH in the Southern California Bight
Authors: Nam SungHyun , Takeshita Yuichiro , Frieder Christina A. , Martz Todd , Ballard John
Published: July 2015

Chemical properties of the California Undercurrent (CU) have been changing over the past several decades, yet the mechanisms responsible for the trend are still not fully understood. We present a survey of temperature, salinity, O2, pH, and currents at intermediate depths (defined here as 50–500 m) in the summer (30 June to 10 July) and winter (8–15 December) of 2012 in the southern region of the Southern California Bight. Observations of temperature, salinity, and currents reveal that local bathymetry and small gyres play an important role in the flow path of the California Undercurrent (CU). Using spiciness (π) as a tracer, we observe a 10% increase of Pacific Equatorial Water (PEW) in the core of the CU during the summer versus the winter. This is associated with an increase in π of 0.2, and a decrease in O2 and pH of 30 μmol kg−1 and 0.022, respectively; the change in pH is driven by increased CO2, while total alkalinity remains unchanged. The high‐π, low‐O2, and low‐pH waters during the summer are not distributed uniformly in the study region. Moreover, mooring observations at the edge of the continental shelf reveal intermittent intrusions of PEW onto the shelf with concomitant decreases in O2 and pH. We estimate that increased advection of PEW in the CU could account for approximately 50% of the observed decrease in O2, and between 49 and 73% of the decrease in pH, over the past three decades.
Transport properties of silicate melts
Authors: Ni Huaiwei , Hui Hejiu , Steinle‐Neumann Gerd
Published: July 2015

A quantitative description of the transport properties,diffusivity,viscosity,electrical, andthermal conductivity, of silicate melts is essential for understanding melting‐related petrologic and geodynamic processes. We here provide a systematic overview on the current knowledge of these properties from experiments and molecular dynamics simulations, their dependence on pressure, temperature, and composition, atomistic processes underlying them, and physical models to describe their variations. We further establish phenomenological and physical links between diffusivity, viscosity, and electrical conductivity that are based on structural rearrangement in the melt. Neutral molecules and network‐modifying cations with low electric field strength displayintrinsic diffusivity, which is controlled by the intrinsic properties (size and valence) of the species. By contrast, oxygen and network formers with high field strength showextrinsic diffusivity, which is more sensitive to extrinsic parameters including temperature (T), pressure (P), and melt composition (X). SimilarT‐P‐Xdependence of diffusivity and electrical conductivity and their quantitative relation reveal the role of intrinsically diffusing species in electrical transport, while viscosity is tied to the extrinsically diffusing species in a similar way. However, the differences in the structural role and mobility of various atomic species diminish with increasing temperature and/or pressure: all transport processes are increasingly coupled, eventually converging to a uniform rate and mechanism. Accurate comprehension of interatomic interactions and melt structure is vital to fully accounting for the compositional dependence of transport properties, and simple polymerization parameters such as nonbridging oxygen per tetrahedrally coordinated cation are inadequate.
The morphology of streams restored for market and nonmarket purposes: Insights from a mixed natural‐social science approach
Authors: Doyle Martin W. , Singh Jai , Lave Rebecca , Robertson Morgan M.
Published: July 2015

We use geomorphic surveys to quantify the differences between restored and nonrestored streams, and the difference between streams restored for market purposes (compensatory mitigation) from those restored for nonmarket programs. We also analyze the social and political‐economic drivers of the stream restoration and mitigation industry using analysis of policy documents and interviews with key personnel including regulators, mitigation bankers, stream designers, and scientists. Restored streams are typically wider and geomorphically more homogenous than nonrestored streams. Streams restored for the mitigation market are typically headwater streams and part of a large, complex of long restored main channels, and many restored tributaries; streams restored for nonmarket purposes are typically shorter and consist of the main channel only. Interviews reveal that designers integrate many influences including economic and regulatory constraints, but traditions of practice have a large influence as well. Thus, social forces shape the morphology of restored streams.
A Highly Elastic and Rapidly Crosslinkable Elastin‐Like Polypeptide‐Based Hydrogel for Biomedical Applications
Authors: Zhang Yi‐Nan , Avery Reginald K. , Vallmajo‐Martin Queralt , Assmann Alexander , Vegh Andrea , Memic Adnan , Olsen Bradley D. , Annabi Nasim , Khademhosseini Ali
Published: July 2015

Elastin‐like polypeptides (ELPs) are promising for biomedical applications due to their unique thermoresponsive and elastic properties. ELP‐based hydrogels have been produced through chemical and enzymatic crosslinking or photocrosslinking of modified ELPs. Herein, a photocrosslinked ELP gel using only canonical amino acids is presented. The inclusion of thiols from a pair of cysteine residues in the ELP sequence allows disulfide bond formation upon exposure to UV light, leading to the formation of a highly elastic hydrogel. The physical properties of the resulting hydrogel such as mechanical properties and swelling behavior can be easily tuned by controlling ELP concentrations. The biocompatibility of the engineered ELP hydrogels is shown in vitro as well as corroborated in vivo with subcutaneous implantation of hydrogels in rats. ELP constructs demonstrate long‐term structural stability in vivo, and early and progressive host integration with no immune response, suggesting their potential for supporting wound repair. Ultimately, functionalized ELPs demonstrate the ability to function as an in vivo hemostatic material over bleeding wounds.
Enhanced Performance of Organic Solar Cells with Increased End Group Dipole Moment in Indacenodithieno[3,2‐b]thiophene‐Based Molecules
Authors: Intemann Jeremy J. , Yao Kai , Ding Feizhi , Xu Yunxiang , Xin Xukai , Li Xiaosong , Jen Alex K.‐Y.
Published: July 2015

Four new molecular donors are reported using a D1‐A‐D2‐A‐D1 structure, where D1 is an oligiothiophene, A is a benzothiadiazole, and D2 is indacenodithieno[3,2‐b]thiophene. The resulting materials provide efficiencies as high as 6.5% in organic solar cells, without the use of solvent additives or thermal/solvent annealing. A strong correlation between the end group (D1‐A) dipole moment and the fill factor (FF), mobility, and loss in the open‐circuit voltage (VOC) is observed. Indacenodithieno[3,2‐b]thiophene‐fluorobenzothiadiazole‐terthiophene (IDTT‐FBT‐3T) possesses the largest end group dipole moment, and in turn, has the highest mobility, FF, and power conversion efficiency in devices. It also has a similarly high VOC (0.95 V) to the other materials (0.93–0.99 V), despite possessing a much higher highest occupied molecular orbital (HOMO) energy level.
A Universal Approach to Ultrasmall Magneto‐Fluorescent Nanohybrids
Authors: Feld Artur , Merkl Jan‐Philip , Kloust Hauke , Flessau Sandra , Schmidtke Christian , Wolter Christopher , Ostermann Johannes , Kampferbeck Michael , Eggers Robin , Mews Alf , Schotten Theo , Weller Horst
Published: July 2015

Seeded emulsion polymerization is a powerful universal method to produce ultrasmall multifunctional magnetic nanohybrids. In a two‐step procedure, iron oxide nanocrystals were initially encapsulated in a polystyrene (PS) shell and subsequently used as beads for a controlled assembly of elongated quantum dots/quantum rods (QDQRs). The synthesis of a continuous PS shell allows the whole construct to be fixed and the composition of the nanohybrid to be tuned. The fluorescence of the QDQRs and magnetism of iron oxide were perfectly preserved, as confirmed by single‐particle investigation, fluorescence decay measurements, and relaxometry. Bio‐functionalization of the hybrids was straightforward, involving copolymerization of appropriate affinity ligands as shown by immunoblot analysis. Additionally, the universality of this method was shown by the embedment of a broad scale of NPs.
Ruthenium‐Catalyzed Monoalkenylation of Aromatic Ketones by Cleavage of Carbon–Heteroatom Bonds with Unconventional Chemoselectivity
Authors: Kondo Hikaru , Akiba Nana , Kochi Takuya , Kakiuchi Fumitoshi
Published: July 2015

Ruthenium‐catalyzed selective monoalkenylation of ortho CO or CN bonds of aromatic ketones was achieved. The reaction allowed the direct comparison of the relative reactivities of the cleavage of different carbon‐heteroatom bonds, thus suggesting an unconventional chemoselectivity, where smaller, more‐electron‐donating groups are more easily cleaved. Selective monofunctionalization of CO bonds in the presence of ortho CH bonds was also achieved.
The Efficacy and Safety of Subcutaneous Clazakizumab in Patients With Moderate‐to‐Severe Rheumatoid Arthritis and an Inadequate Response to Methotrexate: Results From a Multinational, Phase IIb, Randomized, Double‐Blind, Placebo/Active‐Controlled, Dose‐Ranging Study
Authors: Weinblatt Michael E. , Mease Philip , Mysler Eduardo , Takeuchi Tsutomu , Drescher Edit , Berman Alberto , Xing Jun , Zilberstein Moshe , Banerjee Subhashis , Emery Paul
Published: July 2015

Objective

Clazakizumab is a humanized monoclonal antibody that binds to the interleukin‐6 (IL‐6) cytokine. This study was undertaken to evaluate the efficacy and safety of clazakizumab in combination with methotrexate (MTX) or clazakizumab monotherapy versus MTX alone in patients with rheumatoid arthritis (RA) and an inadequate response to MTX.

Methods

In this multinational, phase IIb, randomized, double‐blind, placebo‐controlled, dose‐ranging study, patients were randomized to receive 1) once‐monthly subcutaneous (SC) clazakizumab at 25, 100, or 200 mg plus MTX, 2) once‐monthly SC clazakizumab at 100 mg or 200 mg as monotherapy, or 3) MTX plus placebo (i.e., MTX alone). Adalimumab (40 mg) plus MTX was included as an active reference. The primary end point was the American College of Rheumatology 20% (ACR20) improvement response rate at week 12. Secondary end points included ACR20, ACR50, and ACR70 response rates as well as protocol‐defined remission rates and Health Assessment Questionnaire disability index scores at weeks 12 and 24.

Results

In total, 418 patients were randomized, and baseline characteristics were balanced across the treatment groups. Patients receving clazakizumab had significantly greater ACR20 response rates at week 12 compared with patients receiving MTX alone (76.3%, 73.3%, and 60.0% of patients in the clazakizumab 25, 100, and 200 mg plus MTX groups, respectively, and 55.0% and 61.0% of patients in the clazakizumab 100 and 200 mg monotherapy groups, respectively, versus 39.3% of patients receiving MTX alone; P < 0.05 for all comparisons). At week 24, all clazakizumab groups had higher ACR20, ACR50, and ACR70 response rates and higher remission rates compared with MTX alone. Rates of serious adverse events ranged from 8.3% to 13.6% in the clazakizumab treatment groups, compared with 3.3% in the MTX alone group. Changes in laboratory data were consistent with the pharmacologic effects of IL‐6 blockade.

Conclusion

In patients with RA and an inadequate response to MTX, treatment with clazakizumab in combination with MTX or clazakizumab monotherapy was well tolerated, and patients achieved significant improvements in disease activity, including higher rates of remission, as compared with patients receiving MTX alone.
Role of the Neutrophil Chemorepellent Soluble Dipeptidyl Peptidase IV in Decreasing Inflammation in a Murine Model of Arthritis
Authors: Herlihy Sarah E. , Brown Monica L. , Pilling Darrell , Weeks Brad R. , Myers Linda K. , Gomer Richard H.
Published: July 2015

Objective

To determine whether an intraarticular injection of the neutrophil chemorepellent dipeptidyl peptidase IV (DPPIV; CD26) can attenuate inflammation and decrease the severity of arthritis in a murine model.

Methods

DBA/1 mice were immunized with type II collagen/Freund's complete adjuvant to produce collagen‐induced arthritis (CIA). On day 25 postimmunization, recombinant human DPPIV (rhDPPIV) or phosphate buffered saline was injected intraarticularly, and arthritis severity scores were recorded 3 times per week. The hind legs of mice in both groups were fixed, decalcified, paraffin embedded, and sectioned. Pathologic scores for inflammation and neutrophil infiltration were recorded on a scale of 1–8, and the number of neutrophils was determined by morphometric cell counts. In addition, Mac‐2–positive macrophages and articular damage were assessed using anti–Mac‐2 antibodies and histologic staining, respectively.

Results

Injection of rhDPPIV reduced the mean score of arthritis severity in mice with CIA. DPPIV treatment reduced the overall extent of inflammation and articular damage around the arthritic joint and periarticular tissue, and also decreased neutrophil and macrophage infiltration.

Conclusion

A localized injection of the neutrophil chemorepellent DPPIV reduces inflammation and the severity of the disease in a murine model of arthritis.
Protein Tyrosine Phosphatase Nonreceptor Type 2: An Important Regulator of lnterleukin‐6 Production in Rheumatoid Arthritis Synovial Fibroblasts
Authors: Aradi Borbala , Kato Masaru , Filkova Maria , Karouzakis Emmanuel , Klein Kerstin , Scharl Michael , Kolling Christoph , Michel Beat A. , Gay Renate E. , Buzas Edit I. , Gay Steffen , Jüngel Astrid
Published: July 2015

Objective

To investigate the role of protein tyrosine phosphatase nonreceptor type 2 (PTPN2) in the pathogenesis of rheumatoid arthritis (RA).

Methods

Synovial tissue samples from patients with RA and patients with osteoarthritis (OA) were stained for PTPN2. Synovial fibroblasts were stimulated with tumor necrosis factor (TNF) and interleukin‐1β (IL‐1β), lipopolysaccharide (LPS), TRAIL, or thapsigargin. The expression of PTPN2 in synovial fibroblasts and peripheral blood mononuclear cells (PBMCs) was analyzed by real‐time polymerase chain reaction and Western blotting. Cell death, the release of IL‐6 and IL‐8, and the induction of autophagy were analyzed after PTPN2 silencing. Methylated DNA immunoprecipitation analysis was used to evaluate DNA methylation–regulated gene expression of PTPN2.

Results

PTPN2 was significantly overexpressed in synovial tissue samples from RA patients compared to OA patients. Patients receiving anti‐TNF therapy showed significantly reduced staining for PTPN2 compared with patients treated with nonbiologic agents. PTPN2 expression was higher in RA synovial fibroblasts (RASFs) than in OASFs. This differential expression was not regulated by DNA methylation. PTPN2 was further up‐regulated after stimulation with TNF, TNF combined with IL‐1β, or LPS. There was no significant difference in basal PTPN2 expression in PBMCs from patients with RA, ankylosing spondylitis, or systemic lupus erythematosus or healthy controls. Most interestingly, PTPN2 silencing in RASFs significantly increased the production of the inflammatory cytokine IL‐6 but did not affect levels of IL‐8. Moreover, functional analysis showed that high PTPN2 levels contributed to the increased apoptosis resistance of RASFs and increased autophagy.

Conclusion

This is the first study of PTPN2 in RASFs showing that PTPN2 regulates IL‐6 production, cell death, and autophagy. Our findings indicate that PTPN2 is linked to the pathogenesis of RA via synovial fibroblasts.
Brief Report: Enrichment of Activated Group 3 Innate Lymphoid Cells in Psoriatic Arthritis Synovial Fluid
Authors: Leijten Emmerik F. A. , van Kempen Tessa S. , Boes Marianne , Michels‐van Amelsfort Jocea M. R. , Hijnen Dirkjan , Hartgring Sarita A. Y. , van Roon Joel A. G. , Wenink Mark H. , Radstake Timothy R. D. J.
Published: July 2015

Objective

Innate lymphoid cells (ILCs) are a recently discovered group of cells that are essential to epithelial homeostasis and are implicated in psoriasis pathogenesis, yet they have never been reported in psoriatic arthritis (PsA).

Methods

ILC classes and subsets were characterized in the peripheral blood (PB) of healthy controls, patients with psoriasis, and patients with PsA and in the synovial fluid (SF) of patients with PsA and patients with rheumatoid arthritis (RA). Cell surface marker expression and intracellular cytokine production following stimulation were analyzed using flow cytometry.

Results

ILCs were identified in the SF and were 4‐fold more abundant in PsA SF than in PsA PB. Fewer CCR6+ ILCs were found in PsA PB than in healthy control PB, while PsA SF was enriched for CCR6+ ILCs compared to PsA PB and RA SF. Natural cytotoxicity receptor NKp44+ group 3 ILCs were rare in PB and RA SF, but abundant in PsA SF. Increased numbers of interleukin‐17A (IL‐17A)–producing ILCs were present in PsA SF compared to RA SF. CCR6, NKp44, and melanoma cell adhesion molecule (MCAM) were expressed on the cell surface of SF ILCs that produced IL‐17A. The number of circulating NKp44+, CCR6+, and MCAM+ ILCs in blood was inversely correlated with PsA disease activity.

Conclusion

Our findings indicate that PsA SF is enriched for group 3 ILCs that express CCR6 and NKp44, which distinguishes the synovial compartment from RA. The increased IL‐17A production by SF ILCs indicates a novel role for ILCs in PsA.
Safety and Efficacy of Atacicept in Combination With Rituximab for Reducing the Signs and Symptoms of Rheumatoid Arthritis: A Phase II, Randomized, Double‐Blind, Placebo‐Controlled Pilot Trial
Authors: van Vollenhoven R. F. , Wax S. , Li Y. , Tak P. P.
Published: July 2015

Objective

To explore the safety and tolerability of atacicept in combination with rituximab in patients with active rheumatoid arthritis (RA) receiving rituximab re‐treatment.

Methods

In this randomized, double‐blind, placebo‐controlled pilot trial, 2 infusions (1,000 mg per infusion) of intravenous rituximab, given 2 weeks apart, were followed by once‐weekly subcutaneous injections of 150 mg atacicept or placebo for 25 weeks. Primary end points were the nature, incidence, and severity of adverse events (AEs). Secondary end points were the effects on peripheral blood B cells, disease activity biomarkers, and American College of Rheumatology 20% (ACR20), 50% (ACR50), and 70% (ACR70) response rates.

Results

Eighteen patients were randomized to receive atacicept and 9 to receive placebo. AEs occurred in 17 atacicept‐treated patients (94.4%) and in all 9 placebo‐treated patients (100%). There were no infection‐related serious adverse events. Hypersensitivity and injection site reactions were more common, and more patients withdrew due to AEs, in the atacicept group. Median reductions in Ig levels from baseline to week 32 were greater with atacicept (median change in IgG −31.2%, IgM −60.9%, and IgA −56.4%) than with placebo (median change in IgG −4.4%, IgM −15.9%, and IgA −8.2%). Peripheral B cell numbers remained low in all patients after rituximab‐mediated B cell depletion, limiting comparison of time to recovery between treatment groups. There were no between‐group differences in ACR20, ACR50, and ACR70 response rates.

Conclusion

In this exploratory trial, atacicept in combination with rituximab showed no new safety issues. Peripheral B cell counts remained too low to determine whether atacicept delayed B cell re‐expansion following rituximab‐mediated depletion. Despite clear biologic effects, adding atacicept to rituximab in patients with active RA was not associated with clinical benefit.
Brief Report: Intraarticular Sprifermin Not Only Increases Cartilage Thickness, but Also Reduces Cartilage Loss: Location‐Independent Post Hoc Analysis Using Magnetic Resonance Imaging
Authors: Eckstein Felix , Wirth Wolfgang , Guermazi Ali , Maschek Susanne , Aydemir Aida
Published: July 2015

Objective

To determine whether an anabolic drug (sprifermin) is capable of reducing cartilage loss wherever it occurs in a given knee, using a subject‐specific, location‐independent analysis of cartilage change in patients with knee osteoarthritis (OA).

Methods

Study participants (n = 168; ages ≥40 years, 69% women) had symptomatic femorotibial OA not confined to the medial compartment. Sprifermin (10, 30, or 100 μg) or placebo was injected intraarticularly 3 times over 3 weeks, both after randomization (baseline) and 3 months later. Coronal magnetic resonance images were acquired at baseline and 3, 6, and 12 months after treatment. The femorotibial cartilage of each subject was segmented, and changes in cartilage thickness were computed across 16 subregions. Location‐independent post hoc analysis was used to compute summary scores of negative and positive changes in the subregions, summarized as the total cartilage thinning sum score (ThCTnS) and the total cartilage thickening sum score (ThCTkS), capturing change in either direction in each knee. Ordered values of the magnitude of subject‐specific subregional changes in thickness were determined. The ThCTnS and ThCTkS in each sprifermin dose group at 12 months of followup were compared with the values in the matched placebo groups, using the Wilcoxon‐Mann‐Whitney test.

Results

The mean ± SD ThCTnS was −591 ± 617 μm (median −360 μm, Q1/Q3 = −820/−200 μm) in patients treated with 100 μg sprifermin (n = 57), and −921 ± 777 μm (median −745 μm, Q1/Q3 = −1,190/−380 μm) in patients given placebo (n = 18). The mean difference in the ThCTnS between the 100‐μg sprifermin group and the placebo group was 331 μm (95% confidence interval [95% CI] 24, 685), a difference that was statistically significant (P = 0.03). The mean difference in the ThCTkS in the 100‐μg sprifermin group compared with the placebo group was 237 μm (95% CI 34, 440), also a statistically significant difference (P = 0.028).

Conclusion

Sprifermin not only increases cartilage thickness, but also reduces cartilage loss. Subject‐specific, location‐independent analysis of both cartilage thinning and thickening represents a sensitive and informative approach for studying the effects of disease‐modifying OA drugs.
Role of Bcl‐3 in the Development of Follicular Helper T Cells and in the Pathogenesis of Rheumatoid Arthritis
Authors: Meguro Kazuyuki , Suzuki Kotaro , Hosokawa Junichi , Sanayama Yoshie , Tanaka Shigeru , Furuta Shunsuke , Ikeda Kei , Takatori Hiroaki , Suto Akira , Sakamoto Akemi , Ohara Osamu , Nakajima Hiroshi
Published: July 2015

Objective

We have previously shown that expression of the Bcl‐3 gene, a member of the IκB family, is down‐regulated in CD4+ T cells from patients with rheumatoid arthritis (RA) following tocilizumab therapy. The objective of this study was to examine the role of Bcl‐3 in the pathogenesis of RA.

Methods

DNA microarray analysis was used to compare the signal intensity of Bcl‐3 in CD4+ T cells from untreated RA patients and healthy controls. We examined the roles of interleukin‐6 (IL‐6)/STAT‐3 signaling in the induction of Bcl‐3. In addition, we analyzed the gene expression profiles of Bcl‐3–transduced CD4+ T cells by RNA sequencing. The effects of enforced expression as well as gene silencing of Bcl‐3 on the development of follicular helper T (Tfh) cells were evaluated. Finally, we examined correlations between the signal intensities of Bcl‐3 and Tfh cell–related genes in CD4+ T cells from untreated RA patients.

Results

Bcl‐3 levels were significantly higher in RA patients than in healthy controls. IL‐6 induced Bcl‐3 expression in CD4+ T cells in a STAT‐3–dependent manner. Transcriptome analysis revealed that the expression of Bcl‐6, a master regulator of Tfh cell differentiation, was significantly up‐regulated by the enforced Bcl‐3 expression. The enforced Bcl‐3 expression increased, but Bcl‐3 silencing decreased, the numbers of IL‐21–producing Tfh‐like cells. Bcl‐3 levels in CD4+ T cells from RA patients correlated positively with the levels of Tfh cell–related genes CXCR5, inducible costimulator, and achaete‐scute homolog 2.

Conclusion

Bcl‐3 is involved in the development of Tfh cells and the pathogenesis of RA, presumably by inducing IL‐21 production.
Risk Factors for the Rapid Increase in Risk of Acute Coronary Events in Patients With New‐Onset Rheumatoid Arthritis: A Nested Case–Control Study
Authors: Mantel Ängla , Holmqvist Marie , Nyberg Fredrik , Tornling Göran , Frisell Thomas , Alfredsson Lars , Askling Johan
Published: July 2015

Objective

To investigate risk factors for acute coronary syndrome (ACS) in patients with new‐onset rheumatoid arthritis (RA).

Methods

We performed a nested case–control study of patients with incident RA included in the Epidemiological Investigation of RA study. Cases with ACS were identified using Swedish national health registers and matched with up to 5 controls without ACS, based on incidence density–based sampling. Information on potential exposures (clinical disease activity, serologic features, genetic markers, comorbidities, pharmacotherapies, and sick leave) was collected from medical charts and register‐based sources.

Results

We identified 138 cases and 624 controls. Smoking, history of myocardial infarction, and >50 days of sick leave the year following RA onset were associated with an increased risk of ACS. Area under the curve measurements of C‐reactive protein level, erythrocyte sedimentation rate, Disease Activity Score in 28 joints (DAS28), and global health in the upper tertile during the first year and the complete followup period were both strongly associated with an increased risk of ACS. Treatment with disease‐modifying antirheumatic drugs did not alter the ACS risk, nor did the presence of rheumatoid factor (RF) or shared epitope alleles, whereas high anti–citrullinated protein antibody (ACPA) levels were borderline significantly associated with ACS risk.

Conclusion

In this study of risk factors for ACS in incident RA, clinical markers of inflammatory activity, disease activity, and total number of days of sick leave and disability pension during the first year following RA onset were identified as ACS risk factors. We found no association with RF, which was previously linked to cardiovascular disease risk in RA, but there was a borderline significant association with high ACPA levels.
Asymmetric Total Synthesis of (−)‐Maoecrystal V
Authors: Zhang Wei‐bin , Shao Wen‐bin , Li Fu‐zhuo , Gong Jian‐xian , Yang Zhen
Published: July 2015

The asymmetric total synthesis of (−)‐maoecrystal V, a novel cytotoxic pentacyclic ent‐kaurane diterpene, has been accomplished. Key steps of the current strategy involve an early‐stage semipinacol rearrangement reaction for the construction of the C10 quaternary stereocenter, a rhodium‐catalyzed intramolecular O−H insertion reaction, and a sequential Wessely oxidative dearomatization/intramolecular Diels–Alder reaction to forge the pentacyclic framework of maoecrystal V.
Development of a new procedure for the determination of captopril in pharmaceutical formulations employing chemiluminescence and a multicommuted flow analysis approach
Authors: Lima Manoel J. A. , Fernandes Ridvan N. , Tanaka Auro A. , Reis Boaventura F.
Published: July 2015

This paper describes a new technique for the determination of captopril in pharmaceutical formulations, implemented by employing multicommuted flow analysis. The analytical procedure was based on the reaction between hypochlorite and captopril. The remaining hypochlorite oxidized luminol that generated electromagnetic radiation detected using a homemade luminometer. To the best of our knowledge, this is the first time that this reaction has been exploited for the determination of captopril in pharmaceutical products, offering a clean analytical procedure with minimal reagent usage. The effectiveness of the proposed procedure was confirmed by analyzing a set of pharmaceutical formulations. Application of the paired t‐test showed that there was no significant difference between the data sets at a 95% confidence level. The useful features of the new analytical procedure included a linear response for captopril concentrations in the range 20.0–150.0 µmol/L (r = 0.997), a limit of detection (3σ) of 2.0 µmol/L, a sample throughput of 164 determinations per hour, reagent consumption of 9 µg luminol and 42 µg hypochlorite per determination and generation of 0.63 mL of waste. A relative standard deviation of 1% (n = 6) for a standard solution containing 80 µmol/L captopril was also obtained. Copyright © 2015 John Wiley & Sons, Ltd.