Evaluation of dynamic recurrence risk for locally advanced gastric cancer in the clinical setting of adjuvant chemotherapy: a real-world study with IPTW-based conditional recurrence analysis.

BACKGROUND: The long-term dynamic recurrence hazard of locally advanced gastric cancer (LAGC) in the clinical setting of adjuvant chemotherapy (ACT) remains unclear. PURPOSE: This study aimed to investigate the dynamic recurrence risk of LAGC in patients who received ACT or not. METHODS: The study assessed data from patients with LAGC who underwent radical gastrectomy between January, 2010 and October, 2015. Inverse probability of treatment weighting (IPTW) was performed to reduce selection bias between the ACT and observational (OBS) groups. Conditional recurrence-free survival (cRFS) and restricted mean survival time (RMST) were used to assess the survival differences. RESULTS: In total, 1,661 LAGC patients were included (ACT group, n = 1,236 and OBS group, n = 425). The recurrence hazard gradually declined; in contrast, cRFS increased with RFS already accrued. Following IPTW adjustment, the cRFS rates were higher in the ACT group than those in the OBS group for patients at baseline or with accrued RFS of 1 and 2 years (p˂0.05). However, the cRFS rates of the ACT group were comparable with those of the OBS group for patients with accrued RFS of 3 or more years (p > 0.05). Likewise, the 5-year △RMST between the ACT and OBS groups demonstrated a similar trend. Moreover, the hematological metastasis rate of the ACT group was significantly lower than that of the OBS group for patients at baseline or with accrued RFS of 1 and 2 years, respectively (p˂0.05). CONCLUSIONS: Although ACT could provide substantial benefits for patients with LAGC, the differences in recurrence hazard between the ACT and OBS groups may attenuate over time, which could help guide surveillance and alleviate patients' anxiety. Further prospective large-scale studies are warranted.

Understanding Conformational Dynamics of Complex Lipid Mixtures Relevant to Biology.

This is a perspective article entitled "Frontiers in computational biophysics: understanding conformational dynamics of complex lipid mixtures relevant to biology" which is following a CECAM meeting with the same name.

2D lattice model of a lipid bilayer: Microscopic derivation and thermodynamic exploration.

Based on all-atom Molecular Dynamics (MD) simulations of a lipid bilayer we present a systematic mapping on a 2D lattice model. Keeping the lipid type and the chain order parameter as key variables we derive a free energy functional, containing the enthalpic interaction of adjacent lipids as well as the tail entropy. The functional form of both functions is explicitly determined for saturated and polyunsaturated lipids. By studying the lattice model via Monte Carlo simulations it is possible to reproduce the temperature dependence of the distribution of order parameters of the pure lipids, including the prediction of the gel transition. Furthermore, application to a mixture of saturated and polyunsaturated lipids yields the correct phase separation behavior at lower temperatures with a simulation time reduced by approximately 7 orders of magnitude as compared to the corresponding MD simulations. Even the time-dependence of the de-mixing is reproduced on a semi-quantitative level. Due to the generality of the approach we envisage a large number of further applications, ranging from modeling larger sets of lipids, sterols, and solvent proteins to predicting nucleation barriers for the melting of lipids. Particularly, from the properties of the 2D lattice model one can directly read off the enthalpy and entropy change of the 1,2-dipalmitoyl-sn-glycero-3-phosphocholine gel-to-liquid transition in excellent agreement with experimental and MD results.

Optical torque reversal and spin-orbit rotational Doppler shift experiments.

We report on optical rotational Doppler frequency shift experiments in the context of a counter-intuitive optomechanical phenomenon that is the angular analog of so-called negative optical radiation forces, which involves spin-orbit scattering of light. In practice, spin-orbit opto-mechanical effects arising from the interaction between polarized light and azimuthally varying birefringent optical elements are retrieved from mechano-optical experiments that involve spatial of the medium. Two kinds of experiments (single-beam and two-beam geometries) are performed and both approaches are discussed in the framework of previous dynamical geometric phase and rotational Doppler shift experiments based on spin and/or orbital angular momentum of light.

Molecular dynamics study of interaction and substrate channeling between neuron-specific enolase and B-type phosphoglycerate mutase.

Phosphoglycerate mutase (PGM) and enolase are consecutive enzymes in the glycolytic pathway. We used molecular dynamics simulation to examine the interaction of human B-type PGM (dPGM-B) and neuron-specific enolase (NSE). Specifically, we studied the interactions of 31 orientations of these enzymes by means of the effective energy function implicit solvation method. Interactions between active regions of the enzymes occurred preferentially, although the strongest interactions appeared to be between the back side of NSE and the active regions of dPGM-B. Cleavage of 2PG from dPGM-B was investigated, and the Ser(14)-Leu(30) loop of dPGM-B is suggested as a cleavage site and, likely, another entrance site of a ligand. Substrate channeling between the enzymes was observed when NSE with its active regions Leu(11)-Asn(16), Arg(49)-Lys(59), and Gly(155)-Ala(158) covered the Ser(14)-Leu(30) loop of dPGM-B. Analyses of the results make us believe that the channeling between PGM and enolase "benefits" from weak interaction. The probability of formation of channeling favorable complex is estimated to be up to 5%, while functional interaction between NSE and dPGM-B might be as high as 20%. NSE and dPGM-B functional interaction seems not to be isotype specific.

Laser-Assisted Bioprinting for Bone Repair.

Bioprinting is a novel technological approach that has the potential to solve unmet questions in the field of tissue engineering. Laser-assisted bioprinting (LAB), due to its unprecedented cell printing resolution and precision, is an attractive tool for the in situ printing of a bone substitute. Here, we describe the protocol for LAB and its use for the in situ bioprinting of mesenchymal stromal cells, associated with collagen and nanohydroxyapatite, in order to favor bone regeneration in a calvaria defect model in mice.

Laser-assisted 3D bioprinting of exocrine pancreas spheroid models for cancer initiation study.

Pancreatic ductal adenocarcinoma (PDAC) is the most common malignancy of the pancreas. It has shown a poor prognosis and a rising incidence in the developed world. Other pathologies associated with this tissue include pancreatitis, a risk condition for pancreatic cancer. The onset of both pancreatitis and pancreatic cancer follows a common pattern: exocrine pancreatic acinar cells undergo a transdifferentiation to duct cells that triggers a 3D restructuration of the pancreatic tissue. However, the exact mechanism underlying this process remains partially undefined. Further understanding the cellular events leading to PDAC could open new avenues in the development of novel therapeutic approaches. Since current 2D cell culture models fail to mimic the tridimensional complexity of the pancreatic tissue, new in vitro models are urgently needed. Here, we generated 3D pancreatic cell spheroid arrays using laser-assisted bioprinting and characterized their phenotypic evolution over time through image analysis and phenotypic characterization. We show that these bioprinted spheroids, composed of both acinar and ductal cells, can replicate the initial stages of PDAC development. This bioprinted miniaturized spheroid-based array model should prove useful for the study of the internal and external factors that contribute to the formation of precursor PDAC lesions and to cancer progression, and may therefore shed light on future PDAC therapy strategies.

Comparing an All-Atom and a Coarse-Grained Description of Lipid Bilayers in Terms of Enthalpies and Entropies: From MD Simulations to 2D Lattice Models.

Simulations of lipid bilayers at different levels of detail is of major interest in computational biophysics. With a recently proposed algorithm, by mapping the information from a Molecular Dynamics (MD) simulation of a lipid bilayer on a 2D lattice model, it is possible to gain insight into the enthalpic and entropic contributions, governing the interaction of adjacent lipids and the individual chain entropy, respectively. The contributions are obtained as a function of the lipid chain order parameter. Here we use this approach for the case of pure DPPC (1,2-dipalmitoyl-sn-glycero-3-phosphocholine) to compare the all-atom CHARMM36 DPPC lipid with the respective MARTINI coarse-grained DPPC lipid. We individually compare the enthalpy and entropy functions as well as the resulting free energies. This allows us to gain new insight into the comparison of both levels of description. When numerically solving the 2D lattice model via Monte Carlo (MC) simulations, the 2D model displays the gel/liquid transition at the same temperature as the respective MD simulation. The dramatic increase in efficiency of the 2D lattice model as compared to the MD simulation allows us to estimate the equilibrium order parameters of the gel phase, inaccessible by the MD simulations even after 3 µs.

In situ prevascularization designed by laser-assisted bioprinting: effect on bone regeneration.

Vascularization plays a crucial role in bone formation and regeneration process. Development of a functional vasculature to improve survival and integration of tissue-engineered bone substitutes remains a major challenge. Biofabrication technologies, such as bioprinting, have been introduced as promising alternatives to overcome issues related to lack of prevascularization and poor organization of vascular networks within the bone substitutes. In this context, this study aimed at organizing endothelial cells in situ, in a mouse calvaria bone defect, to generate a prevascularization with a defined architecture, and promote in vivo bone regeneration. Laser-assisted bioprinting (LAB) was used to pattern Red Fluorescent Protein-labeled endothelial cells into a mouse calvaria bone defect of critical size, filled with collagen containing mesenchymal stem cells and vascular endothelial growth factor. LAB technology allowed safe and controlled in vivo printing of different cell patterns. In situ printing of endothelial cells gave rise to organized microvascular networks into bone defects. At two months, vascularization rate (vr) and bone regeneration rate (br) showed statistically significant differences between the 'random seeding' condition and both 'disc' pattern (vr = +203.6%; br = +294.1%) and 'crossed circle' pattern (vr = +355%; br = +602.1%). These results indicate that in vivo LAB is a valuable tool to introduce in situ prevascularization with a defined configuration and promote bone regeneration.

Modeling of annexin A2-Membrane interactions by molecular dynamics simulations.

The annexins are a family of Ca2+-regulated phospholipid binding proteins that are involved in membrane domain organization and membrane trafficking. Although they are widely studied and crystal structures are available for several soluble annexins their mode of membrane association has never been studied at the molecular level. Here we obtained molecular information on the annexin-membrane interaction that could serve as paradigm for the peripheral membrane association of cytosolic proteins by Molecular Dynamics simulations. We analyzed systems containing the monomeric annexin A2 (AnxA2), a membrane with negatively charged phosphatidylserine (POPS) lipids as well as Ca2+ ions. On the atomic level we identify the AnxA2 orientations and the respective residues which display the strongest interaction with Ca2+ ions and the membrane. The simulation results fully agree with earlier experimental findings concerning the positioning of bound Ca2+ ions. Furthermore, we identify for the first time a significant interaction between lysine residues of the protein and POPS lipids that occurs independently of Ca2+ suggesting that AnxA2-membrane interactions can also occur in a low Ca2+ environment. Finally, by varying Ca2+ concentrations and lipid composition in our simulations we observe a calcium-induced negative curvature of the membrane as well as an AnxA2-induced lipid ordering.

Key molecular requirements for raft formation in lipid/cholesterol membranes.

The lipid mixture of DPPC (saturated lipid)/DUPC (unsaturated lipid)/CHOL (cholesterol) is studied with respect to its ability to form liquid-ordered and liquid-disordered phases. We employ coarse-grained simulations with MARTINI force field. All three components are systematically modified in order to explore the relevant molecular properties, leading to phase separation. Specifically, we show that the DPPC/DUPC/CHOL system unmixes due to enthalpic DPPC-DPPC and DPPC-CHOL interactions. The phase separation remains unchanged, except for the formation of a gel phase at long times after decreasing the conformational degrees of freedom of the unsaturated DUPC. In contrast, the phase separation can be suppressed by softening the DPPC chains. In an attempt to mimic the ordering and unmixing effect of CHOL the latter is replaced by a stiff and shortened DPPC-like lipid. One still observes phase separation, suggesting that it is mainly the rigid and planar structure of CHOL which is important for raft formation. Addition of an extra bead to the head of CHOL has no notable impact on the phase separation of the system, supporting the irrelevance of the Umbrella model for the phase separation. Reduction of the conformational entropy of CHOL by stiffening its last bead results in a significant increase of the order of the DPPC/CHOL domain. This suggests that the conformational entropy of CHOL is important to prohibit the gelation process. The interleaflet interactions as mediated by the terminal molecular groups seem to have a strong impact on the possibility of a subsequent gelation process after phase separation.

Phase separation in a lipid/cholesterol system: comparison of coarse-grained and united-atom simulations.

Ternary mixtures of saturated and unsaturated phospholipids and cholesterol constitute a well-known model system to study raft formation in membranes. This phenomenon is, e.g., observed in cell membranes. Here, coarse-grained (CG) and microscopic united-atom (UA) simulations are performed to investigate the phase separation of a membrane bilayer for the ternary system of saturated 16:0 (DPPC) and unsaturated 18:2 (DUPC) phospholipids and cholesterol (CHOL). The results of a 9 µs UA simulation demonstrate the onset of phase separation and can be compared with properties of the corresponding CG system. While sharing the structural features of phase separation in the CG model, the onset of demixing for the UA model is 40 times slower. This factor can be rationalized by the different short-time diffusion constants. Various system properties such as order parameters and lipid-CHOL and CHOL-CHOL interactions are analyzed and compared between the UA and CG models. From the structural perspective, the phase separation process appears to be rather similar for both models, which illustrates once more the power of using CG approaches.