Engineering a Pro-Osteogenic Secretome through the Transient Silencing of the Gene Encoding Secreted Frizzled Related Protein 1.

The evidence sustaining the regenerative properties of mesenchymal stem cells' (MSCs) secretome has prompted a paradigm change, where MSCs have shifted from being considered direct contributors to tissue regeneration toward being seen as cell factories for producing biotech medicines. We have previously designed a method to prime MSCs towards osteogenic differentiation by silencing the Wnt/ß-Catenin inhibitor Sfpr1. This approach produces a significant increase in bone formation in osteoporotic mice. In this current work, we set to investigate the contribution of the secretome from the MSCs where Sfrp1 has been silenced, to the positive effect seen on bone regeneration in vivo. The conditioned media (CM) of the murine MSCs line C3H10T1/2, where Sfrp1 has been transiently silenced (CM-Sfrp1), was found to induce, in vitro, an increase in the osteogenic differentiation of this same cell line, as well as a decrease of the expression of the Wnt inhibitor Dkk1 in murine osteocytes ex vivo. A reduction in the RANKL/OPG ratio was also detected ex vivo, suggesting a negative effect of CM-Sfrp1 on osteoclastogenesis. Moreover, this CM significantly increases the mineralization of human primary MSCs isolated from osteoportotic patients in vitro. Proteomic analysis identified enrichment of proteins involved in osteogenesis within the soluble and vesicular fractions of this secretome. Altogether, we demonstrate the pro-osteogenic potential of the secretome of MSCs primmed in this fashion, suggesting that this is a valid approach to enhance the osteo-regenerative properties of MSCs' secretome.

Diagnostic Reliability of Plain Radiography in Osteonecrosis of the Femoral Head: General Radiological Features Revised.

BACKGROUND AND OBJECTIVES: Osteonecrosis of the femoral head (ONFH) is an incapacitating disease that frequently results in the collapse of the femoral head and secondary osteoarthritis. The diagnosis and staging of this pathology, which usually rely on imaging studies, are challenging. Currently, conventional radiography is the basis of the initial diagnostic assessment. In recent decades, however, radiographs have been considered insensitive to early changes in ONFH and thus, a suboptimal diagnostic tool. Paradoxically, the imaging features of radiographs are often profuse, substantial, and characteristic. This study aimed to elucidate the real limitations of this radiologic tool by assessing the diagnostic reliability of the key radiologic features and staging. METHODS: This was a retrospective study in which radiographs from 28 idiopathic ONFH confirmed cases who underwent hip arthroplasty were analyzed by eight observers who were asked to identify the presence or absence of ONFH universally reported imaging features in AP hip radiographs. RESULTS: Concordance analysis revealed a poor agreement between observers for most of the assessed imaging features. Only the identification of femoral head flattening and osteoarthritis signs exhibited moderate agreement with statistical significance. In contrast, the detection of radiological osteoporosis and the loss of trabeculation showed the lowest reliability, with negative kappa coefficients. CONCLUSION: There is a lack of agreement between qualified observers, even for the identification of the most characteristic ONFH radiographic feature. The reliability of plain radiography for the detection of basic radiological elements is even weaker in the early stages of the disease.

Communication between bone marrow mesenchymal stem cells and multiple myeloma cells: Impact on disease progression.

Multiple myeloma (MM) is a hematological malignancy characterized by the accumulation of immunoglobulin-secreting clonal plasma cells at the bone marrow (BM). The interaction between MM cells and the BM microenvironment, and specifically BM mesenchymal stem cells (BM-MSCs), has a key role in the pathophysiology of this disease. Multiple data support the idea that BM-MSCs not only enhance the proliferation and survival of MM cells but are also involved in the resistance of MM cells to certain drugs, aiding the progression of this hematological tumor. The relation of MM cells with the resident BM-MSCs is a two-way interaction. MM modulate the behavior of BM-MSCs altering their expression profile, proliferation rate, osteogenic potential, and expression of senescence markers. In turn, modified BM-MSCs can produce a set of cytokines that would modulate the BM microenvironment to favor disease progression. The interaction between MM cells and BM-MSCs can be mediated by the secretion of a variety of soluble factors and extracellular vesicles carrying microRNAs, long non-coding RNAs or other molecules. However, the communication between these two types of cells could also involve a direct physical interaction through adhesion molecules or tunneling nanotubes. Thus, understanding the way this communication works and developing strategies to interfere in the process, would preclude the expansion of the MM cells and might offer alternative treatments for this incurable disease.

Active and durable R2MnRuO7 pyrochlores with low Ru content for acidic oxygen evolution.

The production of green hydrogen in water electrolyzers is limited by the oxygen evolution reaction (OER). State-of-the-art electrocatalysts are based on Ir. Ru electrocatalysts are a suitable alternative provided their performance is improved. Here we show that low-Ru-content pyrochlores (R2MnRuO7, R = Y, Tb and Dy) display high activity and durability for the OER in acidic media. Y2MnRuO7 is the most stable catalyst, displaying 1.5 V at 10 mA cm-2 for 40 h, or 5000 cycles up to 1.7 V. Computational and experimental results show that the high performance is owed to Ru sites embedded in RuMnOx surface layers. A water electrolyser with Y2MnRuO7 (with only 0.2 mgRu cm-2) reaches 1 A cm-2 at 1.75 V, remaining stable at 200 mA cm-2 for more than 24 h. These results encourage further investigation on Ru catalysts in which a partial replacement of Ru by inexpensive cations can enhance the OER performance.

Nanoparticle-mediated selective Sfrp-1 silencing enhances bone density in osteoporotic mice.

Osteoporosis (OP) is characterized by a loss in bone mass and mineral density. The stimulation of the canonical Wnt/ß-catenin pathway has been reported to promote bone formation, this pathway is controlled by several regulators as secreted frizzled-related protein-1 (Sfrp-1), antagonist of the pathway. Thus, Sfrp-1 silencing therapies could be suitable for enhancing bone growth. However, the systemic stimulation of Wnt/ß-catenin has been correlated with side effects. This work hypothesizes the administration of lipid-polymer NPs (LPNPs) functionalized with a MSC specific aptamer (Apt) and carrying a SFRP1 silencing GapmeR, could favor bone formation in OP with minimal undesired effects. Suitable SFRP1 GapmeR-loaded Apt-LPNPs (Apt-LPNPs-SFRP1) were administered in osteoporotic mice and their biodistribution, toxicity and bone induction capacity were evaluated. The aptamer functionalization of the NPs modified their biodistribution profile showing a four-fold increase in the bone accumulation and a ten-fold decrease in the hepatic accumulation compared to naked LPNPs. Moreover, the histological evaluation revealed evident changes in bone structure observing a more compact trabecular bone and a cortical bone thickness increase in the Apt-LPNPs-SFRP1 treated mice with no toxic effects. Therefore, these LPNPs showed suitable properties and biodistribution profiles leading to an enhancement on the bone density of osteoporotic mice.

Osteonecrosis of the Femoral Head: A Multidisciplinary Approach in Diagnostic Accuracy.

Osteonecrosis of the Femoral Head (ONFH) is a disabling disease affecting up to 30,000 people yearly in the United States alone. Diagnosis and staging of this pathology are both technically and logistically challenging, usually relying on imaging studies. Even anatomopathological studies, considered the gold standard for identifying ONFH, are not exempt from problems. In addition, the diagnosis is often made by different healthcare specialists, including orthopedic surgeons and radiologists, using different imaging modes, macroscopic features, and stages. Therefore, it is not infrequent to find disagreements between different specialists. The aim of this paper is to clarify the association and accuracy of ONFH diagnosis between healthcare professionals. To this end, femoral head specimens from patients with a diagnosis of ONFH were collected from patients undergoing hip replacement surgery. These samples were later histologically analyzed to establish an ONFH diagnosis. We found that clinico-radiological diagnosis of ONFH evidences a high degree of histological confirmation, thus showing an acceptable diagnostic accuracy. However, when the diagnoses of radiologists and orthopedic surgeons are compared with each other, there is only a moderate agreement. Our results underscore the need to develop an effective diagnosis based on a multidisciplinary approach to enhance currently limited accuracy and reliability.

Intraoperative brain mapping during awake surgery in symptomatic supratentorial cavernomas.

BACKGROUND: Complete resection of symptomatic supratentorial cavernoma (SCA) and removal of the surrounding gliotic area is recommended to minimize the risk of persistent seizures or (re)bleeding. Surgery of SCA located in an eloquent area, can carry out severe postoperative neurological morbidity. We report a study aimed to assess feasibility, extent of resection and outcome after surgical removal of CA by cortico-subcortical intraoperative brain stimulation (ioBS) in the awake patient. METHODS: Six patients diagnosed of symptomatic SCA located on an eloquent area and operated on while awake under local anaesthesia ioBS, were included. Preoperative planning included neuropsychologic assessment of language-related functions, sociocognitive functions and executive functions. Intraoperatively, we recorded the results achieved in the planned neuropsychological tasks when stimulation was applied (cortical and subcortical). Postoperative control 3D MRI was scheduled at 1 month after surgery to calculate extent of resection. Neuropsychological assessment at 6 months after surgery was performed in all cases. RESULTS: Six patients (5 females, 1 male) aged 24-48 years were included in our study. Locations of the lesions were right insular (n=1), left insular (n=1), left temporo-insular (n=1), left temporal (n=2) and left frontal (n=1). In all patients, positive findings were obtained during ioBS. In 5 patients, complete surgical resection was achieved. Two patients had postoperative transient neurological deficits, one case of hemiparesis, one case of dysnomia, both cleared over a 6-month period. Clinical follow-up revealed that all patients experienced complete recovery from preoperative symptoms within a year and five patients with seizures showed marked improvement and eventually quit antiepileptic drugs. Neuropsychological assessment at 6 months provided normal results compared to preoperative baseline in all domains. CONCLUSIONS: Our study suggests that ioBS in the awake surgery of symptomatic SCA located in eloquent areas, allows to increase the rate of complete resection, minimizing postoperative neurological and neuropsychological deficit, and improving postoperative seizures control.

Effective Osteogenic Priming of Mesenchymal Stem Cells through LNA-ASOs-Mediated Sfrp1 Gene Silencing.

Mesenchymal stem cell (MSC) transplantation has emerged as a promising approach for bone regeneration. Importantly, the beneficial effects of MSCs can be improved by modulating the expression levels of specific genes to stimulate MSC osteogenic differentiation. We have previously shown that Smurf1 silencing by using Locked Nucleic Acid-Antisense Oligonucleotides, in combination with a scaffold that sustainably releases low doses of BMP-2, was able to increase the osteogenic potential of MSCs in the presence of BMP-2 doses significantly smaller than those currently used in the clinic. This would potentially allow an important reduction in this protein in MSs-based treatments, and thus of the side effects linked to its administration. We have further improved this system by specifically targeting the Wnt pathway modulator Sfrp1. This approach not only increases MSC bone regeneration efficiency, but is also able to induce osteogenic differentiation in osteoporotic human MSCs, bypassing the need for BMP-2 induction, underscoring the regenerative potential of this system. Achieving successful osteogenesis with the sole use of LNA-ASOs, without the need of administering pro-osteogenic factors such as BMP-2, would not only reduce the cost of treatments, but would also open the possibility of targeting these LNA-ASOs specifically to MSCs in the bone marrow, allowing us to treat systemic bone loss such as that associated with osteoporosis.

Intraoperative brain mapping during awake surgery in symptomatic supratentorial cavernomas.

BACKGROUND: Complete resection of symptomatic supratentorial cavernoma (SCA) and removal of the surrounding gliotic area is recommended to minimize the risk of persistent seizures or (re)bleeding. Surgery of SCA located in an eloquent area, can carry out severe postoperative neurological morbidity. We report a study aimed to assess feasibility, extent of resection and outcome after surgical removal of CA by cortico-subcortical intraoperative brain stimulation (ioBS) in the awake patient. METHODS: Six patients diagnosed of symptomatic SCA located on an eloquent area and operated on while awake under local anaesthesia ioBS, were included. Preoperative planning included neuropsychologic assessment of language-related functions, sociocognitive functions and executive functions. Intraoperatively, we recorded the results achieved in the planned neuropsychological tasks when stimulation was applied (cortical and subcortical). Postoperative control 3D MRI was scheduled at 1 month after surgery to calculate extent of resection. Neuropsychological assessment at 6 months after surgery was performed in all cases. RESULTS: Six patients (5 females, 1 male) aged 24-48 years were included in our study. Locations of the lesions were right insular (n=1), left insular (n=1), left temporo-insular (n=1), left temporal (n=2) and left frontal (n=1). In all patients, positive findings were obtained during ioBS. In 5 patients, complete surgical resection was achieved. Two patients had postoperative transient neurological deficits, one case of hemiparesis, one case of dysnomia, both cleared over a 6-month period. Clinical follow-up revealed that all patients experienced complete recovery from preoperative symptoms within a year and five patients with seizures showed marked improvement and eventually quit antiepileptic drugs. Neuropsychological assessment at 6 months provided normal results compared to preoperative baseline in all domains. CONCLUSIONS: Our study suggests that ioBS in the awake surgery of symptomatic SCA located in eloquent areas, allows to increase the rate of complete resection, minimizing postoperative neurological and neuropsychological deficit, and improving postoperative seizures control.

Proton-pump Inhibitor Response Prediction Using Esophageal microRNAs in Children With Eosinophilic Esophagitis.

OBJECTIVES: Eosinophilic esophagitis (EoE) is a chronic esophageal disease characterized by eosinophilic inflammation. Proton-pump inhibitors (PPI) induce disease remission but no predictive factors of PPI-responsiveness have been identified yet. So, a biomarker must be found to differentiate between responders (PPI-R) and nonresponder patients (PPI-NR) to PPI. Aims were to identify any molecular biomarker that could predict PPI responsiveness and to study molecular remission after PPI therapy. METHODS: This prospective study enrolled 39 controls and 43 pediatric children with EoE from 2 hospitals, and they were treated with esomeprazole for 8 to 12 weeks. After therapy, patients were classified as either PPI-R or PPI-NR. Biopsies were collected and RNA, microRNAs, and proteins were isolated from them, measuring levels by qPCR and Western blot (WB). Also, miRNAs were evaluated in serum. RESULTS: We found several esophageal miRNAs with different expression values between PPI-R and PPI-NR children, which can be used to discriminate them (area under curve = 0.90). No useful serum miRNAs were, however, identified. Also, these miRNAs were dysregulated in responder patients before and after PPI therapy. Moreover, we corroborated in this child population, that PPI-R displayed a significant decrease in eotaxin-3, IL-5, IL-13, periostin, and major basic protein (P < 0.05) and a significant increase in filaggrin levels after PPI treatment (P < 0.01). CONCLUSIONS: Esophageal miRNA levels found are able to discriminate between both PPI-R and PPI-NR at baseline, and before and after treatment in PPI-R, so they could be used as biomarkers. Furthermore, we observed clinical and esophageal molecular restoration in PPI-R patients after PPI therapy.

Mesenchymal stem cells secretome: The cornerstone of cell-free regenerative medicine.

Mesenchymal stem cells (MSCs) are the most frequently used stem cells in clinical trials due to their easy isolation from various adult tissues, their ability of homing to injury sites and their potential to differentiate into multiple cell types. However, the realization that the beneficial effect of MSCs relies mainly on their paracrine action, rather than on their engraftment in the recipient tissue and subsequent differentiation, has opened the way to cell-free therapeutic strategies in regenerative medicine. All the soluble factors and vesicles secreted by MSCs are commonly known as secretome. MSCs secretome has a key role in cell-to-cell communication and has been proven to be an active mediator of immune-modulation and regeneration both in vitro and in vivo. Moreover, the use of secretome has key advantages over cell-based therapies, such as a lower immunogenicity and easy production, handling and storage. Importantly, MSCs can be modulated to alter their secretome composition to better suit specific therapeutic goals, thus, opening a large number of possibilities. Altogether these advantages now place MSCs secretome at the center of an important number of investigations in different clinical contexts, enabling rapid scientific progress in this field.

Enhancing survival, engraftment, and osteogenic potential of mesenchymal stem cells.

Mesenchymal stem cells (MSCs) are promising candidates for bone regeneration therapies due to their plasticity and easiness of sourcing. MSC-based treatments are generally considered a safe procedure, however, the long-term results obtained up to now are far from satisfactory. The main causes of these therapeutic limitations are inefficient homing, engraftment, and osteogenic differentiation. Many studies have proposed modifications to improve MSC engraftment and osteogenic differentiation of the transplanted cells. Several strategies are aimed to improve cell resistance to the hostile microenvironment found in the recipient tissue and increase cell survival after transplantation. These strategies could range from a simple modification of the culture conditions, known as cell-preconditioning, to the genetic modification of the cells to avoid cellular senescence. Many efforts have also been done in order to enhance the osteogenic potential of the transplanted cells and induce bone formation, mainly by the use of bioactive or biomimetic scaffolds, although alternative approaches will also be discussed. This review aims to summarize several of the most recent approaches, providing an up-to-date view of the main developments in MSC-based regenerative techniques.

Pediatric sigmoid volvulus due to Chagas disease.

Exposure and infections by Trypanosoma cruzi are the fourth cause of loss of potential life years between parasitic and infectious diseases. We describe the case of a 11-year-old patient with intestinal occlusion, surgically treated with intestinal volvulus, the surgical specimen is sent to histopathology reporting Chagasic megacolon. The age range of presentation is a challenge in the absence of nonspecific symptoms. There is no pediatric statistical data that define trypanosomiasis in a latent or chronic state and will be diagnosed in adult stages due to the physiopathological alterations that they will present.

La exposición y las infecciones por Trypanosoma cruzi ocupan el cuarto lugar entre las causas de pérdida de años de vida potenciales por enfermedades parasitarias e infecciosas. Se describe el caso de un niño de 11 años, con cuadro de oclusión intestinal, intervenido quirúrgicamente con datos de vólvulo intestinal. La pieza quirúrgica se envió a histopatología, que reportó megacolon chagásico. El rango de edad de presentación es un reto ante la falta de síntomas inespecíficos. No se cuenta con datos estadísticos pediátricos que definan la tripanosomiasis en estado latente o crónico, y estos niños serán diagnosticados en la etapa adulta por las alteraciones fisiopatológicas que presentarán.

Casimir probe based upon metallized high Q SiN nanomembrane resonator.

We present the instrumentation and measurement scheme of a new Casimir force probe that bridges Casimir force measurements at microscale and macroscale. A metallized high Q silicon nitride nanomembrane resonator is employed as a sensitive force probe. The high tensile stress present in the nanomembrane not only enhances the quality factor but also maintains high flatness over large area serving as the bottom electrode in a sphere-plane configuration. A fiber interferometer is used to readout the oscillation of the nanomembrane and a phase-locked loop scheme is applied to track the change of the resonance frequency. Because of the high quality factor of the nanomembrane and the high stability of the setup, a frequency resolution down to 2 × 10(-9) and a corresponding force gradient resolution of 3 µN/m is achieved. Besides sensitive measurement of Casimir force, our measurement technique simultaneously offers Kelvin probe measurement capability that allows in situ imaging of the surface potentials.

Casimir force and in situ surface potential measurements on nanomembranes.

We present Casimir force measurements in a sphere-plate configuration that consists of a high quality nanomembrane resonator and a millimeter sized gold coated sphere. The nanomembrane is fabricated from stoichiometric silicon nitride metallized with gold. A Kelvin probe method is used in situ to image the surface potentials to minimize the distance-dependent residual force. Resonance-enhanced frequency-domain measurements of the nanomembrane motion allow for very high resolution measurements of the Casimir force gradient (down to a force gradient sensitivity of 3 µN/m). Using this technique, the Casimir force in the range of 100 nm to 2 µm is accurately measured. Experimental data thus obtained indicate that the device system in the measured range is best described with the Drude model.

Analysis of short range forces in opto-mechanical devices with a nanogap.

We employ the finite-difference time-domain method to calculate the dominant short range forces in optomechanical devices, Casimir and gradient optical forces. Numerical results are obtained for typical silicon optomechanical devices and are compared to metallic reference structures, taking into account geometric and frequency dispersion of silicon. Our results indicate that although a small gap is desirable for operating optomechanical devices, the Casimir force offsets the gradient force in strongly coupled optomechanical devices, which has to be taken into account in the design of optical force tunable devices.

Coupling mechanics to charge transport in carbon nanotube mechanical resonators.

Nanoelectromechanical resonators have potential applications in sensing, cooling, and mechanical signal processing. An important parameter in these systems is the strength of coupling the resonator motion to charge transport through the device. We investigated the mechanical oscillations of a suspended single-walled carbon nanotube that also acts as a single-electron transistor. The coupling of the mechanical and the charge degrees of freedom is strikingly strong as well as widely tunable (the associated damping rate is approximately 3 x 10(6) Hz). In particular, the coupling is strong enough to drive the oscillations in the nonlinear regime.