bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of the electrical excitation signal parameters on the geometry of an argon-based non-thermal atmospheric pressure plasma jet
Authors: Benabbas Mohamed Tahar , Sahli Salah , Benhamouda Abdallah , Rebiai Saida
Published: June 2015

A non-thermal atmospheric pressure argon plasma jet for medical applications has been generated using a high-voltage pulse generator and a homemade dielectric barrier discharge (DBD) reactor with a cylindrical configuration. A plasma jet of about 6 cm of length has been created in argon gas at atmospheric pressure with an applied peak to peak voltage and a frequency of 10 kV and 50 kHz, respectively. The length and the shape of the created plasma jet were found to be strongly dependent on the electrode setup and the applied voltage and the signal frequency values. The length of the plasma jet increases when the applied voltage and/or its frequency increase, while the diameter at its end is significantly reduced when the applied signal frequency increases. For an applied voltage of 10 kV, the plasma jet diameter decreases from near 5 mm for a frequency of 10 kHz to less than 1 mm at a frequency of 50 kHz. This obtained size of the plasma jet diameter is very useful when the medical treatment must be processed in a reduced space.

PACS 2008

52.50.Dg; 52.70.-m; 52.80.-s
Viral Vectors Encoding Endomorphins and Serine Histogranin Attenuate Neuropathic Pain Symptoms after Spinal Cord Injury in Rats
Authors: Nasirinezhad Farinaz , Gajavelli Shyam , Priddy Blake , Jergova Stanislava , Zadina James , Sagen Jacqueline
Published: June 2015

Background

The treatment of spinal cord injury (SCI)-induced neuropathic pain presents a challenging healthcare problem. The lack of available robust pharmacological treatments underscores the need for novel therapeutic methods and approaches. Due to the complex character of neuropathic pain following SCI, therapies targeting multiple mechanisms may be a better choice for obtaining sufficient long-term pain relief. Previous studies in our ab showed analgesic effects using combinations of an NMDA antagonist peptide [Ser1]histogranin (SHG), and the mu-opioid peptides endomorphins (EMs), in several pain models. As an alternative to drug therapy, this study evaluated the analgesic potential of these peptides when delivered via gene therapy.

Results

Lentiviruses encoding SHG and EM-1 and EM-2 were intraspinally injected, either singly or in combination, into rats with clip compression SCI 2 weeks following injury. Treated animals showed significant reduction in mechanica and thermal hypersensitivity, compared to control groups injected with GFP vector only. The antinociceptive effects of individually injected components were modest, but the combination of EMs and SHG produced robust and sustained antinociception. The onset of the analgesic effects was observed between 1–5 weeks post-injection and sustained without decrement for at least 7 weeks. No adverse effects on locomotor function were observed. The involvement of SHG and EMs in the observed antinociception was confirmed by pharmacologic inhibition using intrathecal injection of either the opioid antagonist naloxone or an anti-SHG antibody. Immunohistochemical analysis showed the presence of SHG and EMs in the spinal cord of treated animals, and immunodot-blot analysis of CSF confirmed the presence of these peptides in injected animals. In a separate group of rats, delayed injection of viral vectors was performed in order to mimic a more likely clinical scenario. Comparable and sustained antinociceptive effects were observed in these animals using the SHG-EMs combination vectors compared to the group with early intervention.

Conclusions

Findings from this study support the potential for direct gene therapy to provide a robust and sustained alleviation of chronic neuropathic pain following SCI. The combination strategy utilizing potent mu-opioid peptides with a naturally-derived NMDA antagonist may produce additive or synergistic analgesic effects without the tolerance development for long-term management of persistent pain.
A comparison of conditional random fields and structured support vector machines for chemical entity recognition in biomedical literature
Authors: Tang Buzhou , Feng Yudong , Wang Xiaolong , Wu Yonghui , Zhang Yaoyun , Jiang Min , Wang Jingqi , Xu Hua
Published: June 2015

Background

Chemical compounds and drugs (together called chemical entities) embedded in scientific articles are crucial for many information extraction tasks in the biomedical domain. However, only a very limited number of chemical entity recognition systems are publically available, probably due to the lack of large manually annotated corpora. To accelerate the development of chemical entity recognition systems, the Spanish National Cancer Research Center (CNIO) and The University of Navarra organized a challenge on Chemical and Drug Named Entity Recognition (CHEMDNER). The CHEMDNER challenge contains two individual subtasks: 1) Chemical Entity Mention recognition (CEM); and 2) Chemical Document Indexing (CDI). Our study proposes machine learning-based systems for the CEM task.

Methods

The 2013 CHEMDNER challenge organizers provided a manually annotated 10,000 UTF8-encoded PubMed abstracts according to a predefined annotation guideline: a training set of 3,500 abstracts, a development set of 3,500 abstracts and a test set of 3,000 abstracts. We developed machine learning-based systems, based on conditional random fields (CRF) and structured support vector machines (SSVM) respectively, for the CEM task for this data set. The effects of three types of word representation (WR) features, generated by Brown clustering, random indexing and skip-gram, on both two machine learning-based systems were also investigated. The performance of our system was evaluated on the test set using scripts provided by the CHEMDNER challenge organizers. Primary evaluation measures were micro Precision, Recall, and F-measure.

Results

Our best system was among the top ranked systems with an official micro F-measure of 85.05%. Fixing a bug caused by inconsistent features marginally improved the performance (micro F-measure of 85.20%) of the system.

Conclusions

The SSVM-based CEM systems outperformed the CRF-based CEM systems when using the same features. Each type of the WR feature was beneficial to the CEM task. Both the CRF-based and SSVM-based systems using the all three types of WR features showed better performance than the systems using only one type of the WR feature.
Intracellular complexes of the early-onset torsion dystonia-associated AAA+ ATPase TorsinA
Authors: Li Hui , Wu Hui-Chuan , Liu Zhonghua , Zacchi Lucia F , Brodsky Jeffrey L , Zolkiewski Michal
Published: June 2015

A single GAG codon deletion in the gene encoding torsinA is linked to most cases of early-onset torsion dystonia. TorsinA is an ER-localized membrane-associated ATPase from the AAA+ superfamily with an unknown biological function. We investigated the formation of oligomeric complexes of torsinA in cultured mammalian cells and found that wild type torsinA associates into a complex with a molecular weight consistent with that of a homohexamer. Interestingly, the dystonia-linked variant torsinAΔE displayed a reduced propensity to form the oligomers compared to the wild type protein. We also discovered that the deletion of the N-terminal membrane-associating region of torsinA abolished oligomer formation. Our results demonstrate that the dystonia-linked mutation in the torsinA gene produces a protein variant that is deficient in maintaining its oligomeric state and suggest that ER membrane association is required to stabilize the torsinA complex.
Rituximab for pulmonary lymphomatoid granulomatosis which developed as a complication of methotrexate and azathioprine therapy for rheumatoid arthritis
Authors: Barakat Athar , Grover Karan , Peshin Rohit
Published: June 2015

We report the case of a patient with Rheumatoid Arthritis [RA] presenting with clinical-pathological and radiological features of Pulmonary Lymphomatoid Granulomatosis (PLG). This is a rare lung disorder characterized by multiple nodular lesions with lymphocytic invasion of vascular walls. We present one such case of PLG secondary to Methotrexate and Azathioprine therapy, who was successfully treated with Steroids and Rituximab. We wish to highlight the importance of lung biopsy in the diagnosis and the use of rituximab as a treatment modality for RA as well as PLG.
Registration system of cloud campus by using android smart tablet
Authors: Kamada Shin , Ichimura Takumi , Shigeyasu Tetsuya , Takemoto Yasuhiko
Published: June 2015

Near Field Communication (NFC) standard covers communication protocols and data exchange formats. NFC technology is one of radio-frequency identification (RFID) standards. In Japan, Felica card is a popular way to identify the unique ID. We developed the attendance management system (AMS) as the Android application which works in the smart tablet with NFC. Generally, the AMS in the university is fixed to the wall and each student touches or slides his/her own card to the dedicated equipment. Because a teacher can use his/her own smart tablet and/or smartphone, the attendance records are viewed anytime and anywhere. Moreover, we developed the collecting system between PC and some tablets by using Android beam. Any personal data are encrypted and the file can be transferred over the NFC Bluetooth Handover between PC Linux and smart tablet. By the mining of the collected records, early discovery for chronic non-attenders are extracted in educational affairs section. In this paper, a registration system on the cloud campus system by using the personal smartphone with NFC is developed. The system enables to introduce the university courses that are open to the general public.
The stigma of epilepsy and its effects on marital status
Authors: Riasi Hamidreza , Rajabpour Sanati Ali , Ghaemi Kazem
Published: June 2015

Background

No previous studies have examined marital status of patients with epilepsy and epilepsy-related factors on perceived and enacted stigmas in Iran. In the present study, marital status of patients with epilepsy (PWE’s) in Birjand city in the east of Iran was investigated.

Methods

A multicenter cross-sectional study was conducted to identify factors contributing to the marital status of PWE’s in a cross-sectional study with 471 participants. Diagnosis of epilepsy in participants (374 cases) was confirmed by at least two neurologists.

Results

Marriage rate of PWE’s was 27.3% (n = 102 patients) and divorce rate was 54.8% (n = 205 patients). Divorce rate in women was significantly higher than in men (62.6% vs. 46.4%; P < 0.05), and there were no significant differences between the different types of epilepsy (P > 0.05).

Conclusions

The stigma of epilepsy has impacts on marital status of PWE’s. The PWE’s suffering from the enacted stigma of epilepsy are significantly more likely to get divorced in comparison with other patients.
The McDonald exponentiated gamma distribution and its statistical properties
Authors: Al-Babtain Abdulhakim A , Merovci Faton , Elbatal Ibrahim
Published: June 2015

In this paper, we propose a five-parameter lifetime model called the McDonald exponentiated gamma distribution to extend beta exponentiated gamma, Kumaraswamy exponentiated gamma and exponentiated gamma, among several other models. We provide a comprehensive mathematical treatment of this distribution. We derive the moment generating function and the rth moment. We discuss estimation of the parameters by maximum likelihood and provide the information matrix.

AMS Subject Classification

Primary 62N05; secondary 90B25
Fast sequences MR imaging at the investigation of painful skeletal sites in patients with hip osteonecrosis
Authors: Zibis Aristidis H , Varitimidis Sokratis E , Dailiana Zoe H , Karantanas Apostolos H , Arvanitis Dimitrios L , Malizos Konstantinos N
Published: June 2015

Background

Multiple osteonecrotic foci can be clinically silent when located in metaphyses and becomes painful when it affects juxta-articular areas. The purpose of this study was to assess the value of fast MR imaging to depict the underlying pathology in cases with skeletal pain other than the already diagnosed hip osteonecrosis.

Methods/design

Between 2008 and 2013, 49 patients with already diagnosed hip osteonecrosis reported symptoms of deep skeletal pain in an anatomical site different from the affected hip joint. All patients after thorough history & clinical examination underwent evaluation with x-rays and a single fat suppressed sequence with MR Imaging applying either T2-w TSE or STIR-TSE at the painful site. False positive and false negative findings were recorded for the conventional x-rays and compared to MRI.

Discussion

Forty four (89.8%) patients were positive for osteonecrotic lesions in this study and 76 symptomatic osteonecrosis lesions were revealed at 14 distinct anatomic sites. The agreement between the x-ray findings and the MR imaging regarding osteonecrosis was 46.9%. Plain x-rays showed 43.4% sensitivity, 100% specificity, 100% positive predictive value and 10.4% negative predictive value.

Fast MR imaging with fat suppressed sequences is necessary and adequate as a single method for the investigation of painful skeletal sites in patients with already diagnosed hip osteonecrosis. It allows early diagnosis of the potentially debilitating multiple juxta-articular lesions and consequently their prompt management.
Performance of a novel KRAS mutation assay for formalin-fixed paraffin embedded tissues of colorectal cancer
Authors: Sakai Kazuko , Yoneshige Azusa , Ito Akihiko , Ueda Yoji , Kondo Satoshi , Nobumasa Hitoshi , Fujita Yoshihiko , Togashi Yosuke , Terashima Masato , De Velasco Marco A , Tomida Shuta , Nishio Kazuto
Published: June 2015

We compared the performance of the 3D-Gene® mutation assay (3D-Gene® KRAS mutation assay kit) with the Scorpion-ARMS (therascreen® KRAS RGQ PCR Kit) and Luminex (MEBGEN™ KRAS kit) assays for the detection of KRAS mutations in formalin-fixed, paraffin-embedded tissue samples from 150 patients diagnosed with colorectal cancer. DNA was extracted from the paraffin-embedded tissue samples with or without macrodissection under hematoxylin and eosin staining and the KRAS mutation status was independently determined using these assays. Discordant results were re-analyzed by Sanger sequencing. Mutation detection analysis was successfully performed in all 150 specimens using the 3D-Gene® mutation assay without an invalid case. The concordance rate between the 3D-Gene® mutation assay and Scorpion-ARMS or Luminex was 98.7% (148/150). KRAS mutations were detected at a frequency of 35.3% (53/150) in colorectal cancer specimens. Three discrepant cases were found between the three assays. Overall, our results demonstrate a high concordance rate of between the 3D-Gene® mutation assay and the two existing in-vitro diagnostics kits. All three assays proved to be validated methods for detecting clinically significant KRAS mutations in paraffin-embedded tissue samples.
Testing the tunnel effect: comparison, age and happiness in UK and German panels
Authors: FitzRoy Felix R , Nolan Michael A , Steinhardt Max F , Ulph David
Published: June 2015

In contrast to previous results combining all ages, we findpositiveeffects of comparison income on happiness for the under 45s and negative effects for those over 45. In the UK, these coefficients are several times the magnitude of own income effects. In West Germany, they cancel out to givenoeffect of comparison income on life satisfaction in the whole sample when controlling for fixed effects, time-in-panel, and age-groupings. Pooled OLS estimation gives the usual negative comparison effect in the whole sample for both West Germany and the UK. The residual age-happiness relationship is hump-shaped in all three countries. Results are consistent with a simple life cycle model of relative income under uncertainty.

Jel codes

D10, I31, J10
Preparation and Transport Performances of High-Density, Aligned Carbon Nanotube Membranes
Authors: Zhang Lei , Zhao Bin , Jiang Chuan , Yang Junhe , Zheng Guangping
Published: June 2015

We report a simple and effective method for the preparation of high-density and aligned carbon nanotube (CNT) membranes. The CNT arrays were prepared by water-assisted chemical vapor deposition (CVD) and were subsequently pushed over and stacked into dense membranes by mechanical rolling. It was demonstrated that various gases and liquids, including H2, He, N2, O2, Ar, water, ethanol, hexane, and kerosene, could effectively pass through the aligned carbon nanotube membranes. The membranes exhibited different selections on different gases, indicating that there was a separation potential for the gas mixtures. The selectivities (H2 relative to other gases) of H2/He, H2/N2, H2/O2, and H2/Ar were found to be lower than that of the ideal Knudsen model. For pure water, the permeability was measured to be 3.23 ± 0.05 ml·min−1·cm−2 at 1 atm, indicating that the CNT membranes were promising for applications in liquid filtration and separation.
Extensive rewiring of epithelial-stromal co-expression networks in breast cancer
Authors: Oh Eun-Yeong , Christensen Stephen M , Ghanta Sindhu , Jeong Jong Cheol , Bucur Octavian , Glass Benjamin , Montaser-Kouhsari Laleh , Knoblauch Nicholas W , Bertos Nicholas , Saleh Sadiq MI , Haibe-Kains Benjamin , Park Morag , Beck Andrew H
Published: June 2015

Background

Epithelial-stromal crosstalk plays a critical role in invasive breast cancer pathogenesis; however, little is known on a systems level about how epithelial-stromal interactions evolve during carcinogenesis.

Results

We develop a framework for building genome-wide epithelial-stromal co-expression networks composed of pairwise co-expression relationships between mRNA levels of genes expressed in the epithelium and stroma across a population of patients. We apply this method to laser capture micro-dissection expression profiling datasets in the setting of breast carcinogenesis. Our analysis shows that epithelial-stromal co-expression networks undergo extensive rewiring during carcinogenesis, with the emergence of distinct network hubs in normal breast, and estrogen receptor-positive and estrogen receptor-negative invasive breast cancer, and the emergence of distinct patterns of functional network enrichment. In contrast to normal breast, the strongest epithelial-stromal co-expression relationships in invasive breast cancer mostly represent self-loops, in which the same gene is co-expressed in epithelial and stromal regions. We validate this observation using an independent laser capture micro-dissection dataset and confirm that self-loop interactions are significantly increased in cancer by performing computational image analysis of epithelial and stromal protein expression using images from the Human Protein Atlas.

Conclusions

Epithelial-stromal co-expression network analysis represents a new approach for systems-level analyses of spatially localized transcriptomic data. The analysis provides new biological insights into the rewiring of epithelial-stromal co-expression networks and the emergence of epithelial-stromal co-expression self-loops in breast cancer. The approach may facilitate the development of new diagnostics and therapeutics targeting epithelial-stromal interactions in cancer.
quantro: a data-driven approach to guide the choice of an appropriate normalization method
Authors: Hicks Stephanie C. , Irizarry Rafael A.
Published: June 2015

Normalization is an essential step in the analysis of high-throughput data. Multi-sample global normalization methods, such as quantile normalization, have been successfully used to remove technical variation. However, these methods rely on the assumption that observed global changes across samples are due to unwanted technical variability. Applying global normalization methods has the potential to remove biologically driven variation. Currently, it is up to the subject matter experts to determine if the stated assumptions are appropriate. Here, we propose a data-driven alternative. We demonstrate the utility of our method (quantro) through examples and simulations. A software implementation is available from http://www.bioconductor.org/packages/release/bioc/html/quantro.html.
Scalable microfluidics for single-cell RNA printing and sequencing
Authors: Bose Sayantan , Wan Zhenmao , Carr Ambrose , Rizvi Abbas H. , Vieira Gregory , Pe’er Dana , Sims Peter A.
Published: June 2015

Many important biological questions demand single-cell transcriptomics on a large scale. Hence, new tools are urgently needed for efficient, inexpensive manipulation of RNA from individual cells. We report a simple platform for trapping single-cell lysates in sealed, picoliter microwells capable of printing RNA on glass or capturing RNA on beads. We then develop a scalable technology for genome-wide, single-cell RNA-Seq. Our device generates pooled libraries from hundreds of individual cells with consumable costs of $0.10–$0.20 per cell and includes five lanes for simultaneous experiments. We anticipate that this system will serve as a general platform for single-cell imaging and sequencing.
Impact of autoimmune risk alleles on the immune system
Authors: Ray John P , Hacohen Nir
Published: June 2015

Genetic analyses of autoimmune diseases have revealed hundreds of disease-associated DNA variants, but the identity and function of the causal variants are understudied and warrant deeper mechanistic studies. Here, we highlight methods for deciphering how alleles that are associated with autoimmune disease alter the human immune system, and suggest strategies for future autoimmune genetic research.
Perceived barriers in the use of ultrasound in developing countries
Authors: Shah Sachita , Bellows Blaise A. , Adedipe Adeyinka A. , Totten Jodie E. , Backlund Brandon H. , Sajed Dana
Published: June 2015

Background

Access to ultrasound has increased significantly in resource-limited settings, including the developing world; however, there remains a lack of sonography education and ultrasound-trained physician support in developing countries. To further investigate this potential knowledge gap, our primary objective was to assess perceived barriers to ultrasound use in resource-limited settings by surveying care providers who practice in low- and middle-income settings.

Methods

A 25-question online survey was made available to health care providers who work with an ultrasound machine in low- and middle-income countries (LMICs), including doctors, nurses, technicians, and clinical officers. This was a convenience sample obtained from list-serves of ultrasound and radiologic societies. The survey was analyzed, and descriptive results were obtained.

Results

One hundred and thirty-eight respondents representing 44 LMICs including countries from the continents of Africa, South America, and Asia completed the survey, with a response rate of 9.6 %. Ninety-one percent of the respondents were doctors, and 9 % were nurses or other providers. Applications for ultrasound were diverse, including obstetrics (75 %), DVT evaluation (51 %), abscess evaluation (54 %), cardiac evaluation (64 %), inferior vena cava (IVC) assessment (49 %), Focused Assessment Sonography for Trauma (FAST) exam (64 %), biliary tree assessment (54 %), and other applications. The respondents identified the following barriers to use of ultrasound: lack of training (60 %), lack of equipment (45 %), ultrasound machine malfunction (37 %), and lack of ultrasound maintenance capability (47 %). Seventy-four percent of the respondents wished to have further training in ultrasound, and 82 % were open to receiving distance learning or telesonography training. Subjects used communication tools including Skype, Dropbox, emailed photos, and picture archiving and communication system (PACS) as ways to communicate and receive feedback on ultrasound images.

Conclusions

Health care providers in the developing world identify lack of training as a primary barrier to regular use of ultrasound in their practice. While equipment requirements including maintenance and cost of machines are also important factors, future research is warranted on best practices for training methods, including telesonography and distance learning to enhance ultrasound use in low-resource settings.
The role of beneficial bacteria wall elasticity in regulating innate immune response
Authors: Мokrozub Viktoria V. , Lazarenko Liudmyla M. , Sichel Liubov M. , Babenko Lidia P. , Lytvyn Petro M. , Demchenko Olga M. , Melnichenko Yulia O. , Boyko Nadiya V. , Biavati Bruno , DiGioia Diana , Bubnov Rostyslav V. , Spivak Mykola Ya
Published: June 2015

Background

Probiotics have great potential to contribute to development of healthy dietary regimes, preventive care, and an integrated approach to immunity-related disease management. The bacterial wall is a dynamic entity, depending on many components and playing an essential role in modulating immune response. The impact of cell wall elasticity on the beneficial effects of probiotic strains has not been sufficiently studied.

The aim was to investigate the effect of lactic acid bacteria (LAB) and bifidobacteria strains on phagocytic system cells (macrophages) as related to bacterial wall elasticity, estimated using atomic force microscopy (AFM).

Methods

We conducted studies on Balb/c line mice 18–20 g in weight using lyophilized strains of LAB—Lactobacillus acidophilus IMV B-7279, Lactobacillus casei IMV B-7280, Lactobacillus delbrueckii subsp. bulgaricus IMV B-7281, and bifidobacteria—Bifidobacterium animalis VKL and Bifidobacterium animalis VKB. We cultivated the macrophages obtained from the peritoneal cavity of mice individually with the strains of LAB and bifidobacteria and evaluated their effect on macrophages, oxygen-dependent bactericidal activity, nitric oxide production, and immunoregulatory cytokines. We used AFM scanning to estimate bacterial cell wall elasticity.

Results

All strains had a stimulating effect on the functional activity of macrophages and ability to produce NO/NO2 in vitro. Lactobacilli strains increased the production of IL-12 and IFN-γ in vitro. The AFM demonstrated different cell wall elasticity levels in various strains of LAB and bifidobacteria. The rigidity of the cell walls among lactobacilli was distributed as follows: Lactobacillus acidophilus IMV B-7279 > Lactobacillus casei IMV B-7280 > Lactobacillus delbrueckii subsp. bulgaricus IMV B-7281; among the strains of bifidobacteria: B. animalis VKB > B. animalis VKL. Probiotic strain survival in the macrophages depended on the bacterial cell wall elasticity and on the time of their joint cultivation.

Conclusion

LAB and bifidobacteria strains stimulate immune-modulatory cytokines and active oxygen and nitrogen oxide compound production in macrophages. Strains with a more elastic cell wall according to AFM data demonstrated higher resistance to intracellular digestion in macrophages and higher level of their activation.

AFM might be considered as a fast and accurate method to assess parameters of probiotic strain cell wall to predict their immune-modulatory properties.
The role and potential of umbilical cord blood in an era of new therapies: a review
Authors: Roura Santiago , Pujal Josep-Maria , Gálvez-Montón Carolina , Bayes-Genis Antoni
Published: June 2015

In light of pioneering findings in the 1980s and an estimation of more than 130 million global annual births, umbilical cord blood (UCB) is considered to be the most plentiful reservoir of cells and to have regenerative potential for many clinical applications. Although UCB is used mainly against blood disorders, the spectrum of diseases for which it provides effective therapy has been expanded to include non-hematopoietic conditions; UCB has also been used as source for regenerative cell therapy and immune modulation. Thus, collection and banking of UCB-derived cells have become a popular option. However, there are questions regarding the cost versus the benefits of UCB banking, and it also raises complex ethical and legal issues. This review discusses many issues surrounding the conservation of UCB-derived cells and the great potential and current clinical applications of UCB in an era of new therapies. In particular, we describe the practical issues inherent in UCB collection, processing, and long-term storage as well as the different types of ‘stem’ or progenitor cells circulating in UCB and their uses in multiple clinical settings. Given these considerations, the trend toward UCB will continue to provide growing assistance to health care worldwide.
A new application of cell-free bone regeneration: immobilizing stem cells from human exfoliated deciduous teeth-conditioned medium onto titanium implants using atmospheric pressure plasma treatment
Authors: Omori Masahiro , Tsuchiya Shuhei , Hara Kenji , Kuroda Kensuke , Hibi Hideharu , Okido Masazumi , Ueda Minoru
Published: June 2015

Introduction

Surface modification of titanium (Ti) implants promotes bone formation and shortens the osseointegration period. The aim of this study was to promote bone regeneration and stability around implants using atmospheric pressure plasma (APP) pretreatment. This was followed by immobilization of stem cells from human exfoliated deciduous teeth-conditioned medium (SHED-CM) on the Ti implant surface.

Methods

Ti samples (implants, discs, powder) were treated with APP for 30 seconds. Subsequently, these were immobilized on the treated Ti surface, soaked and agitated in phosphate-buffered saline or SHED-CM for 24 hours at 37 °C. The surface topography of the Ti implants was observed using scanning electron microscopy with energy dispersive X-ray spectroscopy. In vivo experiments using Ti implants placed on canine femur bone were then conducted to permit histological analysis at the bone-implant boundary. For the in vitro experiments, protein assays (SDS-PAGE, Bradford assay, liquid chromatography-ion trap mass spectrometry) and canine bone marrow stromal cell (cBMSC) attachment assays were performed using Ti discs or powder.

Results

In the in vitro study, treatment of Ti implant surfaces with SHED-CM led to calcium phosphate and extracellular matrix protein immobilization. APP pretreatment increased the amount of SHED-CM immobilized on Ti powder, and contributed to increased cBMSC attachment on Ti discs. In the in vivo study, histological analysis revealed that the Ti implants treated with APP and SHED-CM stimulated new bone formation around implants.

Conclusions

Implant device APP pretreatment followed by SHED-CM immobilization may be an effective application to facilitate bone regeneration around dental implants.
What quantitative mechanical loading stimulates in vitro cultivation best?
Authors: Natenstedt Jerry , Kok Aimee C , Dankelman Jenny , Tuijthof Gabrielle JM
Published: June 2015

Articular cartilage has limited regeneration capacities. One of the factors that appear to affect the in vitro cultivation of articular cartilage is mechanical stimulation. So far, no combination of parameters has been identified that offers the best results. The goal is to review the literature in search of the best available set of quantitative mechanical stimuli that lead to optimal in vitro cultivation.

The databases Scopus and PubMed were used to survey the literature, and strict in- and exclusion criteria were applied regarding the presence of quantitative data. The review was performed by studying the type of loading (hydrostatic compression or direct compression), the loading magnitude, the frequency and the loading regime (duration of the loading) in comparison to quantitative evidence of cartilage quality response (cellular, signaling and mechanical).

Thirty-three studies met all criteria of which 8 studied human, 20 bovine, 2 equine, 1 ovine, 1 porcine and 1 canine cells using four different types of cultivated constructs. Six studies investigated loading magnitude within the same setup, three studies the frequency, and seven the loading regime. Nine studies presented mechanical tissue response. The studies suggest that a certain threshold exits for enhanced cartilage in vitro cultivation of explants (>20 % strain and 0.5 Hz), and that chondrocyte-seeded cultivated constructs show best results when loaded with physiological mechanical stimuli. That is a loading pressure between 5–10 MPa and a loading frequency of 1 Hz exerted at intermittent intervals for a period of a week or longer. Critical aspects remain to be answered for translation into in vivo therapies.
Relative genomic stability of adipose tissue derived mesenchymal stem cells: analysis of ploidy, H19 long non-coding RNA and p53 activity
Authors: Ravid Orly , Shoshani Ofer , Sela Meirav , Weinstock Ada , Sadan Tommy Weiss , Gur Eyal , Zipori Dov , Shani Nir
Published: June 2015

Introduction

Mesenchymal stem cells (MSCs) are multipotent and have been derived from various tissues. Although MSCs share many basic features, they often display subtle tissue specific differences. We previously demonstrated that bone marrow (BM) MSCs frequently become polyploid in culture. This tendency was mediated by a reduction in the expression of H19 long non-coding RNA during the transition from a diploid to a polyploid state.

Methods

MSCs were derived from both BM and adipose tissue of mice and expanded under normoxic and hypoxic culture conditions. Cells were stained by propidium iodide and their ploidy was evaluated by FACS. Gene expression of independent MSC preparations was compared by quantitative real time PCR and protein expression levels by Western blot analysis. p53 silencing in MSCs was performed by a specific small hairpin RNA (shRNA).

Results

We set to examine whether genomic instability is common to MSCs originating from different tissues. It is demonstrated that adipose derived MSCs (ASCs) tend to remain diploid during culture while a vast majority of BM MSCs become polyploid. The diploid phenotype of ASCs is correlated with reduced H19 expression compared to BM MSCs. Under hypoxic conditions (3% oxygen) both ASCs and BM MSCs demonstrate increased RNA expression of H19 and Vascular endothelial growth factor A. Importantly, ASC gene expression is significantly less variable than BM MSCs under both oxygen conditions, indicating to their superior homogeneity. Gene expression analysis revealed that p53 target genes, often induced by DNA damage, are up-regulated in ASCs under basal conditions. However, p53 activation following treatment with DNA damaging agents was strongly elevated in BM MSCs compared to ASCs. We found that p53 is involved in maintaining the stable diploid state of ASCs as p53 shRNA induced ploidy changes in ASCs but not in BM MSCs.

Conclusions

The increased genomic stability of murine ASCs together with their lower H19 expression and relative homogeneity suggest a tissue specific higher stability of ASCs compared to BM MSCs, possibly due to higher activity of p53. The tissue specific differences between MSCs from a different tissue source may have important consequences on the use of various MSCs both in vitro and in vivo.
Human mesenchymal stem cells/multipotent stromal cells consume accumulated autophagosomes early in differentiation
Authors: Nuschke Austin , Rodrigues Melanie , Stolz Donna B , Chu Charleen T , Griffith Linda , Wells Alan
Published: June 2015

Introduction

Bone marrow mesenchymal stem cells/multipotent stromal cells (MSCs) are recruited to sites of injury and subsequently support regeneration through differentiation or paracrine activity. During periods of stress such as wound site implant or differentiation, MSCs are subjected to a variety of stressors that might activate pathways to improve cell survival and generate energy. In this study, we monitored MSC autophagy in response to the process of differentiation.

Methods

MSC autophagosome structures were observed by using transmission electron microscopy and a tandem green fluorescent protein-red fluorescent protein autophagic flux reporter to monitor the mammalian microtubule-associated protein-1 light chain 3 (LC3) turnover in real time. MSCs were differentiated by using standard osteogenic and adipogenic media, and autophagy was examined during short-term and long-term differentiation via immunoblots for LC3I and II. Autophagy was modulated during differentiation by using rapamycin and bafilomycin treatments to disrupt the autophagosome balance during the early stages of the differentiation process, and differentiation was monitored in the long term by using Von Kossa and Oil Red O staining as well as quantitative polymerase chain reaction analysis of typical differentiation markers.

Results

We found that undifferentiated MSCs showed an accumulation of a large number of undegraded autophagic vacuoles, with little autophagic turnover. Stimulation of autophagy with rapamycin led to rapid degradation of these autophagosomes and greatly increased rough endoplasmic reticulum size. Upon induction of osteogenic differentiation, MSC expression of LC3II, a common autophagosome marker, was lost within 12 hours, consistent with increased turnover. However, during adipogenic differentiation, drug treatment to alter the autophagosome balance during early differentiation led to changes in differentiation efficiency, with inhibited adipocyte formation following rapamycin treatment and accelerated fat accumulation following autophagosome blockade by bafilomycin.

Conclusions

Our findings suggest that MSCs exist in a state of arrested autophagy with high autophagosome accumulation and are poised to rapidly undergo autophagic degradation. This phenotype is highly sensitive, and a balance of autophagy appears to be key in efficient MSC differentiation and function, as evidenced by our results implicating autophagic flux in early osteogenesis and adipogenesis.
Human adipose-derived stromal/stem cells demonstrate short-lived persistence after implantation in both an immunocompetent and an immunocompromised murine model
Authors: Agrawal Hitesh , Shang Hulan , Sattah Anna Parker , Yang Ning , Peirce Shayn M , Katz Adam J
Published: June 2015

Introduction

Mesenchymal cells are emerging as a promising cell platform for regenerative therapies. However, the fate of cells after transplantation in many different disease settings and tissue beds remains unclear.

Methods

In this study, human adipose-derived stromal/stem (ASCs) cells were fluorescently labeled with a membrane dye and injected into both immunocompetent and immunocompromised mouse strains. Cells were injected either as single cell suspensions, or as self-assembling spheroids. In parallel, cells were purposefully devitalized prior to injection and then implanted in the opposite side in a randomized fashion. These ‘control’ groups were included to determine whether the fluorescent membrane dye would remain localized at the injection site despite the use of nonviable cells. Cell implants and the surrounding tissues were harvested on days 3, 10 and 21 after in vivo delivery and evaluated in a blinded manner. Injection sites were analyzed by fluorescent microscopy, and human cell numbers were quantified using PCR detection of a human-specific endogenous retrovirus (ERV-3). Host response was evaluated by immunofluorescent staining of macrophages.

Results

ERV-3 quantification showed that 95% of the human cells that were viable when they were injected were undetectable at the three-week time-point. Although fluorescent signal persisted for the entire study period, further analysis revealed that much of this signal was located within host macrophages.

Conclusions

These results suggest that human ASCs survive for less than three weeks after injection into even immunocompromised mice, and call into question the notion that human ASCs are immuno-privileged and capable of surviving for extended periods in xenogeneic and/or allogeneic models.
Adipose tissue derived stem cells: in vitro and in vivo analysis of a standard and three commercially available cell-assisted lipotransfer techniques
Authors: Domenis Rossana , Lazzaro Lara , Calabrese Sarah , Mangoni Damiano , Gallelli Annarita , Bourkoula Evgenia , Manini Ivana , Bergamin Natascha , Toffoletto Barbara , Beltrami Carlo A , Beltrami Antonio P , Cesselli Daniela , Parodi Pier Camillo
Published: June 2015

Introduction

Autologous fat grafting is commonly used to correct soft-tissue contour deformities. However, results are impaired by a variable and unpredictable resorption rate. Autologous adipose-derived stromal cells in combination with lipoinjection (cell-assisted lipotransfer) seem to favor a long-term persistence of fat grafts, thus fostering the development of devices to be used in the operating room at the point of care, to isolate the stromal vascular fraction (SVF) and produce SVF-enhanced fat grafts with safe and standardized protocols. Focusing on patients undergoing breast reconstruction by lipostructure, we analyzed a standard technique, a modification of the Coleman’s procedure, and three different commercially available devices (Lipokit, Cytori, Fastem), in terms of 1) ability to enrich fat grafts in stem cells and 2) clinical outcome at 6 and 12 months.

Methods

To evaluate the ability to enrich stem cells, we compared, for each patient (n = 20), the standard lipoaspirate with the respective stem cell-enriched one, analyzing yield, immunophenotype and colony-forming capacity of the SVF cells as well as immunophenotype, clonogenicity and multipotency of the obtained adipose stem cells (ASCs). Regarding the clinical outcome, we compared, by ultrasonography imaging, changes at 6 and 12 months in the subcutaneous thickness of patients treated with stem-cell enriched (n = 14) and standard lipoaspirates (n = 16).

Results

Both methods relying on the enzymatic isolation of primitive cells led to significant increase in the frequency, in the fat grafts, of SVF cells as well as of clonogenic and multipotent ASCs, while the enrichment was less prominent for the device based on the mechanical isolation of the SVF. From a clinical point of view, patients treated with SVF-enhanced fat grafts demonstrated, at six months, a significant superior gain of thickness of both the central and superior-medial quadrants with respect to patients treated with standard lipotransfer. In the median-median quadrant the effect was still persistent at 12 months, confirming an advantage of lipotransfer technique in enriching improving long-term fat grafts.

Conclusions

This comparative study, based on reproducible biological and clinical parameters and endpoints, showed an advantage of lipotransfer technique in enriching fat grafts in stem cells and in favoring, clinically, long-term fat grafts.
Effect of human bone marrow mesenchymal stromal cells on cytokine production by peripheral blood naive, memory, and effector T cells
Authors: Laranjeira Paula , Pedrosa Monia , Pedreiro Susana , Gomes Joana , Martinho Antonio , Antunes Brigida , Ribeiro Tania , Santos Francisco , Trindade Helder , Paiva Artur
Published: June 2015

Introduction

The different distribution of T cells among activation/differentiation stages in immune disorders may condition the outcome of mesenchymal stromal cell (MSC)-based therapies. Indeed, the effect of MSCs in the different functional compartments of T cells is not completely elucidated.

Methods

We investigated the effect of human bone marrow MSCs on naturally occurring peripheral blood functional compartments of CD4+ and CD8+ T cells: naive, central memory, effector memory, and effector compartments. For that, mononuclear cells (MNCs) stimulated with phorbol myristate acetate (PMA) plus ionomycin were cultured in the absence/presence of MSCs. The percentage of cells expressing tumor necrosis factor-alpha (TNF-α), interferon gamma (IFNγ), and interleukin-2 (IL-2), IL-17, IL-9, and IL-6 and the amount of cytokine produced were assessed by flow cytometry. mRNA levels of IL-4, IL-10, transforming growth factor-beta (TGF-β), and cytotoxic T-lymphocyte-associated protein 4 (CTLA4) in purified CD4+ and CD8+ T cells, and phenotypic and mRNA expression changes induced by PMA + ionomycin stimulation in MSCs, were also evaluated.

Results

MSCs induced the reduction of the percentage of CD4+ and CD8+ T cells producing TNF-α, IFNγ, and IL-2 in all functional compartments, except for naive IFNγ+CD4+ T cells. This inhibitory effect differentially affected CD4+ and CD8+ T cells as well as the T-cell functional compartments; remarkably, different cytokines showed distinct patterns of inhibition regarding both the percentage of producing cells and the amount of cytokine produced. Likewise, the percentages of IL-17+, IL-17+TNF-α+, and IL-9+ within CD4+ and CD8+ T cells and of IL-6+CD4+ T cells were decreased in MNC-MSC co-cultures. MSCs decreased IL-10 and increased IL-4 mRNA expression in stimulated CD4+ and CD8+ T cells, whereas TGF-β was reduced in CD8+ and augmented in CD4+ T cells, with no changes for CTLA4. Finally, PMA + ionomycin stimulation did not induce significant alterations on MSCs phenotype but did increase indoleamine-2,3-dioxygenase (IDO), inducible costimulatory ligand (ICOSL), IL-1β, IL-8, and TNF-α mRNA expression.

Conclusions

Overall, our study showed that MSCs differentially regulate the functional compartments of CD4+ and CD8+ T cells, which may differentially impact their therapeutic effect in immune disorders. Furthermore, the influence of MSCs on IL-9 expression can open new possibilities for MSC-based therapy in allergic diseases.
A therapy-grade protocol for differentiation of pluripotent stem cells into mesenchymal stem cells using platelet lysate as supplement
Authors: Luzzani Carlos , Neiman Gabriel , Garate Ximena , Questa María , Solari Claudia , Fernandez Espinosa Darío , García Marcela , Errecalde Ana Lía , Guberman Alejandra , Scassa María Elida , Sevlever Gustavo Emilio , Romorini Leonardo , Miriuka Santiago Gabriel
Published: June 2015

Introduction

Mesenchymal stem cells (MSCs) are a promising source of cells for regenerative therapies. Although they can be isolated easily from several tissues, cell expansion is limited since their properties are lost with successive passages. Hence, pluripotent derived MSCs (PD-MSCs) arise as a suitable alternative for MSC production. Nevertheless, at present, PD-MSC derivation protocols are either expensive or not suitable for clinical purposes.

Methods

In this work we present a therapy-grade, inexpensive and simple protocol to derive MSCs from pluripotent stem cells (PSCs) based on the use of platelet lysate (PL) as medium supplement.

Results

We showed that the PD-MSCPL expressed multiple MSC markers, including CD90, CD73, CD105, CD166, and CD271, among others. These cells also show multilineage differentiation ability and immunomodulatory effects on pre-stimulated lymphocytes. Thorough characterization of these cells showed that a PD-MSCPL resembles an umbilical cord (UC) MSC and differs from a PSC in surface marker and extracellular matrix proteins and integrin expression. Moreover, the OCT-4 promoter is re-methylated with mesenchymal differentiation comparable with the methylation levels of UC-MSCs and fibroblasts. Lastly, the use of PL-supplemented medium generates significantly more MSCs than the use of fetal bovine serum.

Conclusions

This protocol can be used to generate a large amount of PD-MSCs with low cost and is compatible with clinical therapies.