Ex vivo expansion in a clinical grade medium, containing growth factors from human platelets, enhances migration capacity of adipose stem cells.

Introduction: Adipose tissue mesenchymal stem/stromal cells (ASC) can be used as advanced therapy medicinal product in regenerative and cancer medicine. We previously demonstrated Supernatant Rich in Growth Factors (SRGF) can replace fetal bovine serum (FBS) to expand ASC by a clinical grade compliant protocol. The therapeutic potential of ASC is based also on their homing capacity toward inflammatory/cancer sites: oriented cell migration is a fundamental process in this scenario. We investigated the impact of SRGF on ASC migration properties. Methods: The motility/migration potential of ASC expanded in 5% SRGF was analyzed, in comparison to 10% FBS, by standard wound healing, bidimensional chemotaxis and transwell assays, and by millifluidic transwell tests. Mechanisms involved in the migration process were investigated by transient protein overexpression. Results: In comparison to standard 10% FBS, supplementation of the cell culture medium with 5% SRGF, strongly increased migration properties of ASC along the chemotactic gradient and toward cancer cell derived soluble factors, both in static and millifluidic conditions. We showed that, independently from applied migratory stimulus, SRGF expanded ASC were characterized by far lower expression of α-smooth muscle actin (αSMA), a protein involved in the cell migration machinery. Overexpression of αSMA induced a significant and marked decrease in migration capacity of SRGF expanded ASC. Discussion: In conclusion, 5% SRGF addition in the cell culture medium increases the migration potential of ASC, reasonably through appropriate downregulation of αSMA. Thus, SRGF could potentially improve the therapeutic impact of ASC, both as modulators of the immune microenviroment or as targeted drug delivery vehicles in oncology.

Effects of cavity nonlinearities and linear losses on silicon microring-based reservoir computing.

Microring resonators (MRRs) are promising devices for time-delay photonic reservoir computing, but the impact of the different physical effects taking place in the MRRs on the reservoir computing performance is yet to be fully understood. We numerically analyze the impact of linear losses as well as thermo-optic and free-carrier effects relaxation times on the prediction error of the time-series task NARMA-10. We demonstrate the existence of three regions, defined by the input power and the frequency detuning between the optical source and the microring resonance, that reveal the cavity transition from linear to nonlinear regimes. One of these regions offers very low error in time-series prediction under relatively low input power and number of nodes while the other regions either lack nonlinearity or become unstable. This study provides insight into the design of the MRR and the optimization of its physical properties for improving the prediction performance of time-delay reservoir computing.

Native collagen VI delays early muscle stem cell differentiation.

Adult muscle stem cells (MuSCs) are critical for muscle homeostasis and regeneration, and their behavior relies on a finely regulated niche made of specific extracellular matrix (ECM) components and soluble factors. Among ECM proteins, collagen VI (Col6) influences the mechanical properties of the niche and, in turn, MuSC self-renewal capabilities. Here, we investigated whether Col6 can exert a direct function as a biochemical signal for regulating the stemness and differentiation of murine MuSCs and myoblasts. Native Col6, but not its pepsin-resistant fragment, counteracts the early differentiation of myogenic cells by reducing the expression of differentiation marker genes and preserving stemness features, with inhibition of the canonical Wnt pathway. Our data indicate that extracellular Col6 acts as a soluble ligand in delaying early myogenic differentiation by regulating intracellular signals involved in adult myogenesis.

Modeling Doxorubicin Pharmacokinetics in Multiple Myeloma Suggests Mechanism of Drug Resistance.

OBJECTIVE: Multiple myeloma (MM) is a plasma cell malignancy often treated with chemotherapy drugs. Among these, doxorubicin (DOXO) is commonly employed, sometimes in combined-drug therapies, but it has to be optimally administered in order to maximize its efficacy and reduce possible side effects. To support DOXO studies and treatment optimization, here we propose an experimental/modeling approach to establish a model describing DOXO pharmacokinetics (PK) in MM cells. METHODS: A series of in vitro experiments were performed in MM1R and MOLP-2 cells. DOXO was administered at two dosages (200 nM, 450 nM) at [Formula: see text] = 0 and removed at [Formula: see text] = 3 hrs. Intracellular DOXO concentration was measured via fluorescence microscopy during both drug uptake ([Formula: see text] = 0-3 hrs) and release phases ([Formula: see text] = 3-8 hrs). Four PK candidate models were identified, and were compared and selected based on their ability to describe DOXO data and numerical parameter identification. RESULTS: The most parsimonious model consists of three compartments describing DOXO distribution between the extracellular space, the cell cytoplasm and the nucleus, and defines the intracellular DOXO efflux rate through a Hill function, simulating a threshold/saturation drug resistance mechanism. This model predicted DOXO data well in all the experiments and provided precise parameter estimates (mean ± standard deviation coefficient of variation: 15.8 ± 12.2%). CONCLUSIONS: A reliable PK model describing DOXO uptake and release in MM cells has been successfully developed. SIGNIFICANCE: The proposed PK model, once integrated with DOXO pharmacodynamics, has the potential of allowing the study and the optimization of DOXO treatment strategies in MM.

Addressing data scarcity in optical matrix multiplier modeling using transfer learning.

We present and experimentally evaluate the use of transfer learning to address experimental data scarcity when training neural network (NN) models for Mach-Zehnder interferometer mesh-based optical matrix multipliers. Our approach involves pretraining the model using synthetic data generated from a less accurate analytical model and fine-tuning it with experimental data. Our investigation demonstrates that this method yields significant reductions in modeling errors compared to using an analytical model or a standalone NN model when training data is limited. Utilizing regularization techniques and ensemble averaging, we achieve <1 dB root-mean-square error on the 3×3 matrix weights implemented by a photonic chip while using only 25% of the available data.

IRE1a-Induced FilaminA Phosphorylation Enhances Migration of Mesenchymal Stem Cells Derived from Multiple Myeloma Patients.

Multiple myeloma (MM) is an aggressive malignancy that shapes, during its progression, a pro-tumor microenvironment characterized by altered protein secretion and the gene expression of mesenchymal stem cells (MSCs). In turn, MSCs from MM patients can exert an high pro-tumor activity and play a strong immunosuppressive role. Here, we show, for the first time, greater cell mobility paralleled by the activation of FilaminA (FLNA) in MM-derived MSCs, when compared to healthy donor (HD)-derived MSCs. Moreover, we suggest the possible involvement of the IRE1a-FLNA axis in the control of the MSC migration process. In this way, IRE1a can be considered as a good target candidate for MM therapy, considering its pro-survival, pro-osteoclast and chemoresistance role in the MM microenvironment. Our results suggest that IRE1a downregulation could also interfere with the response of MSCs to MM stimuli, possibly preventing cell-cell adhesion-mediated drug resistance. In addition, further investigations harnessing IRE1a-FLNA interaction could improve the homing efficiency of MSC as cell product for advanced therapy applications.

Collagen VI sustains cell stemness and chemotherapy resistance in glioblastoma.

Microenvironmental factors are known fundamental regulators of the phenotype and aggressiveness of glioblastoma (GBM), the most lethal brain tumor, characterized by fast progression and marked resistance to treatments. In this context, the extracellular matrix (ECM) is known to heavily influence the behavior of cancer cells from several origins, contributing to stem cell niches, influencing tumor invasiveness and response to chemotherapy, mediating survival signaling cascades, and modulating inflammatory cell recruitment. Here, we show that collagen VI (COL6), an ECM protein widely expressed in both normal and pathological tissues, has a distinctive distribution within the GBM mass, strongly correlated with the most aggressive and phenotypically immature cells. Our data demonstrate that COL6 sustains the stem-like properties of GBM cells and supports the maintenance of an aggressive transcriptional program promoting cancer cell proliferation and survival. In particular, we identified a specific subset of COL6-transcriptionally co-regulated genes, required for the response of cells to replicative stress and DNA damage, supporting the concept that COL6 is an essential stimulus for the activation of GBM cell response and resistance to chemotherapy, through the ATM/ATR axis. Altogether, these findings indicate that COL6 plays a pivotal role in GBM tumor biology, exerting a pleiotropic action across different GBM hallmarks, including phenotypic identity and gene transcription, as well as response to treatments, thus providing valuable information for the understanding of the complex microenvironmental cues underlying GBM malignancy.

Experimental validation of machine-learning based spectral-spatial power evolution shaping using Raman amplifiers.

We experimentally validate a real-time machine learning framework, capable of controlling the pump power values of Raman amplifiers to shape the signal power evolution in two-dimensions (2D): frequency and fiber distance. In our setup, power values of four first-order counter-propagating pumps are optimized to achieve the desired 2D power profile. The pump power optimization framework includes a convolutional neural network (CNN) followed by differential evolution (DE) technique, applied online to the amplifier setup to automatically achieve the target 2D power profiles. The results on achievable 2D profiles show that the framework is able to guarantee very low maximum absolute error (MAE) (<0.5 dB) between the obtained and the target 2D profiles. Moreover, the framework is tested in a multi-objective design scenario where the goal is to achieve the 2D profiles with flat gain levels at the end of the span, jointly with minimum spectral excursion over the entire fiber length. In this case, the experimental results assert that for 2D profiles with the target flat gain levels, the DE obtains less than 1 dB maximum gain deviation, when the setup is not physically limited in the pump power values. The simulation results also prove that with enough pump power available, better gain deviation (less than 0.6 dB) for higher target gain levels is achievable.

Modified mesenchymal stem cells in cancer therapy: A smart weapon requiring upgrades for wider clinical applications.

Mesenchymal stem stromal cells (MSC) are characterized by the intriguing capacity to home toward cancer cells after systemic administration. Thus, MSC can be harnessed as targeted delivery vehicles of cytotoxic agents against tumors. In cancer patients, MSC based advanced cellular therapies were shown to be safe but their clinical efficacy was limited. Indeed, the amount of systemically infused MSC actually homing to human cancer masses is insufficient to reduce tumor growth. Moreover, induction of an unequivocal anticancer cytotoxic phenotype in expanded MSC is necessary to achieve significant therapeutic efficacy. Ex vivo cell modifications are, thus, required to improve anti-cancer properties of MSC. MSC based cellular therapy products must be handled in compliance with good manufacturing practice (GMP) guidelines. In the present review we include MSC-improving manipulation approaches that, even though actually tested at preclinical level, could be compatible with GMP guidelines. In particular, we describe possible approaches to improve MSC homing on cancer, including genetic engineering, membrane modification and cytokine priming. Similarly, we discuss appropriate modalities aimed at inducing a marked cytotoxic phenotype in expanded MSC by direct chemotherapeutic drug loading or by genetic methods. In conclusion, we suggest that, to configure MSC as a powerful weapon against cancer, combinations of clinical grade compatible modification protocols that are currently selected, should be introduced in the final product. Highly standardized cancer clinical trials are required to test the efficacy of ameliorated MSC based cell therapies.

Emilin-2 is a component of bone marrow extracellular matrix regulating mesenchymal stem cell differentiation and hematopoietic progenitors.

BACKGROUND: Dissection of mechanisms involved in the regulation of bone marrow microenvironment through cell-cell and cell-matrix contacts is essential for the detailed understanding of processes underlying bone marrow activities both under physiological conditions and in hematologic malignancies. Here we describe Emilin-2 as an abundant extracellular matrix component of bone marrow stroma. METHODS: Immunodetection of Emilin-2 was performed in bone marrow sections of mice from 30 days to 6 months of age. Emilin-2 expression was monitored in vitro in primary and mesenchymal stem cell lines under undifferentiated and adipogenic conditions. Hematopoietic stem cells and progenitors in bone marrow of 3- to 10-month-old wild-type and Emilin-2 null mice were analyzed by flow cytometry. RESULTS: Emilin-2 is deposited in bone marrow extracellular matrix in an age-dependent manner, forming a meshwork that extends from compact bone boundaries to the central trabecular regions. Emilin-2 is expressed and secreted by both primary and immortalized bone marrow mesenchymal stem cells, exerting an inhibitory action in adipogenic differentiation. In vivo Emilin-2 deficiency impairs the frequency of hematopoietic stem/progenitor cells in bone marrow during aging. CONCLUSION: Our data provide new insights in the contribution of bone marrow extracellular matrix microenvironment in the regulation of stem cell niches and hematopoietic progenitor differentiation.

Modeling Pharmacokinetics of Doxorubicin in Multiple Myeloma Cells.

Doxorubicin (DOXO) is a well-established chemotherapy drug for treatment of different tumors, ranging from breast cancer, melanoma to multiple myeloma (MM). Here, we present a coupled experimental/modeling approach to study DOXO pharmacokinetics in MM cells, investigate its distribution among the extracellular and intracellular compartments during time. Three model candidates are considered and identified. Model selection is performed based on its ability to describe the data both qualitatively and in terms of quantitative indexes. The most parsimonious model consists of a nonlinear structure with a saturation-threshold control of intracellular DOXO efflux by the DOXO bound to the cellular DNA. This structure could explain the hypothesis that MM cells are drug-resistant, likely due to the involvement of P-glycoproteins.The proposed model is able to predict the intracellular (free and bound) DOXO and suggests the presence of a saturation-threshold drug-resistant mechanism.Clinical Relevance- The model can be used to properly understand and guide further experimental setup, e.g., to investigate multiple myeloma cell variability among different cell lines.

miR-335-laden B Cell-Derived Extracellular Vesicles Promote SOX4-Dependent Apoptosis in Human Multiple Myeloma Cells.

Multiple myeloma (MM) is characterized by the accumulation of malignant plasma cells in the bone marrow. Despite novel therapies, MM still remains an incurable cancer and new strategies are needed. Increased expression of the transcription factor Sex-determining region Y-related high-mobility-group box transcription factor 4 (SOX4) has been correlated with tumor development and progression through a variety of distinct processes, including inhibition of apoptosis, increased cell invasion and metastasis, and induction and maintenance of cancer-initiating cells. The role of SOX4 in MM is largely unknown. Since SOX4 is a known target of miR-335, we used miR-335 to assess whether SOX4 modulation could promote apoptosis in MM cells. Using an MM cell model we show that miR-335 acts both on SOX4-related genes (AKT, PI3K) and hypoxia-inducible factor 1-alpha (Hif1-α). In addition, we show miR-335-laden extracellular vesicles induced in B cells (iEVs) are also effective in targeting SOX4, causing apoptosis. Collectively, we propose that miR-335-laden iEVs could be developed as a novel form of gene therapy in MM.

Nucleofection of Adipose Mesenchymal Stem/Stromal Cells: Improved Transfection Efficiency for GMP Grade Applications.

Nucleofection (NF) is a safe, non-viral transfection method, compatible with Good Manufacturing Practice guidelines. Such a technique is useful to improve therapeutic effectiveness of adipose tissue mesenchymal stem cells (ASC) in clinical settings, but improvement of NF efficiency is mandatory. Supernatant rich in growth factors (SRGF) is a clinical-grade medium additive for ASC expansion. We showed a dramatically increased NF efficiency and post-transfection viability in ASC expanded in presence of SRGF (vs. fetal bovine serum). SRGF expanded ASC were characterized by increased vesicle endocytosis but lower phagocytosis properties. SRGF increased n-6/n-3 ratio, reduced membrane lipid raft occurrence, and lowered intracellular actin content in ASC. A statistical correlation between NF efficiency and lipid raft availability on cell membranes was shown, even though a direct relationship could not be demonstrated: attempts to selectively modulate lipid rafts levels were, in fact, limited by technical constraints. In conclusion, we reported for the first time that tuning clinical-grade compatible cell culture conditions can significantly improve ASC transfection efficiency by a non-viral and safe approach. A deep mechanistic characterization is extremely complex, but we can hypothesize that integrated changes in membrane structure and intracellular actin content could contribute to explain SRGF impact on ASC NF efficiency.

Optimization of frequency combs spectral-flatness using evolutionary algorithm.

We demonstrate the use of meta-heuristics algorithms for flatness optimization of optical frequency combs (OFCs). Without any additional component for flatness compensation, the laser alone is explored when driven by optimized bias current and radio frequency (RF) driving signals composed by multiple harmonics. The bias current amplitude and RF harmonic amplitudes and relative phases are optimized using particle swarm optimization (PSO) and differential evolution (DE) algorithms. The numerical results lead to a 9 lines-GS-laser-based OFC spectrum with 2.9 dB flatness. An online experimental optimization using the DE algorithm results in a 7-line-GS-laser-based OFC with 2 dB flatness.

Inverse design of a Raman amplifier in frequency and distance domains using convolutional neural networks.

We present a convolutional neural network architecture for inverse Raman amplifier design. This model aims at finding the pump powers and wavelengths required for a target signal power evolution in both distance along the fiber and in frequency. Using the proposed framework, the prediction of the pump configuration required to achieve a target power profile is demonstrated numerically with high accuracy in C-band considering both counter-propagating and bidirectional pumping schemes. For a distributed Raman amplifier based on a 100 km single-mode fiber, a low mean set (0.51, 0.54, and 0.64 dB) and standard deviation set (0.62, 0.43, and 0.38 dB) of the maximum test error are obtained numerically employing two and three counter-, and four bidirectional propagating pumps, respectively.

Simultaneous gain profile design and noise figure prediction for Raman amplifiers using machine learning.

A machine learning framework predicting pump powers and noise figure profile for a target distributed Raman amplifier gain profile is experimentally demonstrated. We employ a single-layer neural network to learn the mapping from the gain profiles to the pump powers and noise figures. The obtained results show highly accurate gain profile designs and noise figure predictions, with a maximum error on average of ∼0.3dB. This framework provides a comprehensive characterization of the Raman amplifier and thus is a valuable tool for predicting the performance of next-generation optical communication systems, expected to employ Raman amplification.

Electrospun Structures Made of a Hydrolyzed Keratin-Based Biomaterial for Development of in vitro Tissue Models.

The aim of this study is the analysis and characterization of a hydrolyzed keratin-based biomaterial and its processing using electrospinning technology to develop in vitro tissue models. This biomaterial, extracted from poultry feathers, was mixed with type A porcine gelatin and cross-linked with γ-glycidyloxy-propyl-trimethoxy-silane (GPTMS) to be casted initially in the form of film and characterized in terms of swelling, contact angle, mechanical properties, and surface charge density. After these chemical-physical characterizations, electrospun nanofibers structures were manufactured and their mechanical properties were evaluated. Finally, cell response was analyzed by testing the efficacy of keratin-based structures in sustaining cell vitality and proliferation over 4 days of human epithelial, rat neuronal and human primary skin fibroblast cells.

All-optical OFDM demultiplexing with optical partial Fourier transform and coherent sampling.

We propose a novel scheme with a "time-lens"-based partial optical Fourier transform (OFT) and coherent sampling for high-speed complex orthogonal frequency-division multiplexing (OFDM) signal detection. Compared with all-optical OFDM demultiplexing with a matched optical filter, our proposed method replaces specialized optical filters with commercially available equipment, which relaxes stringent manufacturing and operational requirements. Our simulation shows that even with a partial OFT, theoretically, close to inter-channel interference-free performance is possible. In addition, we performed a proof-of-concept experiment of 16×10 Gbaud quadrature phase-shift keying (QPSK) all-optical OFDM detection, with all the bit error rates far below the 7% hard-overhead forward error correction limit.

Loss of EMILIN-1 Enhances Arteriolar Myogenic Tone Through TGF-ß (Transforming Growth Factor-ß)-Dependent Transactivation of EGFR (Epidermal Growth Factor Receptor) and Is Relevant for Hypertension in Mice and Humans.

Objective- EMILIN-1 (elastin microfibrils interface located protein-1) protein inhibits pro-TGF-ß (transforming growth factor-ß) proteolysis and limits TGF-ß bioavailability in vascular extracellular matrix. Emilin1-/- null mice display increased vascular TGF-ß signaling and are hypertensive. Because EMILIN-1 is expressed in vessels from embryonic life to adulthood, we aimed at unravelling whether the hypertensive phenotype of Emilin1-/- null mice results from a developmental defect or lack of homeostatic role in the adult. Approach and Results- By using a conditional gene targeting inactivating EMILIN-1 in smooth muscle cells of adult mice, we show that increased blood pressure in mice with selective smooth muscle cell ablation of EMILIN-1 depends on enhanced myogenic tone. Mechanistically, we unveil that higher TGF-ß signaling in smooth muscle cells stimulates HB-EGF (heparin-binding epidermal growth factor) expression and subsequent transactivation of EGFR (epidermal growth factor receptor). With increasing intraluminal pressure in resistance arteries, the cross talk established by TGF-ß and EGFR signals recruits TRPC6 (TRP [transient receptor potential] classical type 6) and TRPM4 (TRP melastatin type 4) channels, lastly stimulating voltage-dependent calcium channels and potentiating myogenic tone. We found reduced EMILIN-1 and enhanced myogenic tone, dependent on increased TGF-ß-EGFR signaling, in resistance arteries from hypertensive patients. Conclusions- Taken together, our findings implicate an unexpected role of the TGF-ß-EGFR pathway in hypertension with current translational perspectives.

12 mode, WDM, MIMO-free orbital angular momentum transmission.

Simultaneous MIMO-free transmission of 12 orbital angular momentum (OAM) modes over a 1.2 km air-core fiber is demonstrated. WDM compatibility of the system is shown by using 60, 25 GHz spaced WDM channels with 10 GBaud QPSK signals. System performance is evaluated by measuring bit error rates, which are found to be below the soft FEC limit, and limited by inter-modal crosstalk. The crosstalk in the system is analyzed, and it is concluded that it can be significantly reduced with an improved multiplexer and de-multiplexer.