Identifying low cancer-specific mortality risk lymph node-positive radical prostatectomy patients.

OBJECTIVES: To identify low cancer-specific mortality (CSM) risk lymph node-positive (pN1) radical prostatectomy (RP) patients. METHODS: Within Surveillance, Epidemiology and End Results database (2010-2015) pN1 RP patients were identified. Kaplan-Meier plots and multivariable Cox-regression (MCR) models were used. Pathological characteristics were used to identify patients at lowest CSM risk. RESULTS: Overall, 2197 pN1 RP patients were identified. Overall, 5-year cancer-specific survival (CSS) rate was 93.3%. In MCR models ISUP GG1-2 (hazard ratio [HR]: 0.12, p < 0.001), GG3 (HR: 0.14, p < 0.001), GG4 (HR: 0.35, p = 0.002), pT2 (HR: 0.27, p = 0.012), pT3a (HR: 0.28, p = 0.003), pT3b (HR: 0.39, p = 0.009), and 1-2 positive lymph nodes (HR: 0.64, p = 0.04) independently predicted lower CSM. Pathological characteristics subgroups with the most protective hazard ratios were used to identify low-risk (ISUP GG1-3 and pT2-3a and 1-2 positive lymph nodes) patients versus others (ISUP GG4-5 or pT3b-4 or ≥3 positive lymph nodes). In Kaplan-Meier analyses, 5-year CSS rates were 99.3% for low-risk (n = 480, 21.8%) versus 91.8% (p < 0.001) for others (n = 1717, 78.2%). CONCLUSIONS: Lymph node-positive RP patients exhibit variable CSS rates. Within this heterogeneous group, those at very low risk of CSM may be identified based on pathological characteristics, namely ISUP GG1-3, pT2-3a, and 1-2 positive lymph nodes. Such stratification scheme might be of value for individual patients counseling, as well as in design of clinical trials.

Assessment of the VENUSS and GRANT Models for Individual Prediction of Cancer-specific Survival in Surgically Treated Nonmetastatic Papillary Renal Cell Carcinoma.

Background: Guidelines recommend VENUSS and GRANT models for the prediction of cancer control outcomes after nephrectomy for nonmetastatic papillary renal cell carcinoma (pRCC). Objective: To test the ability of VENUSS and GRANT models to predict 5-yr cancer-specific survival in a North American population. Design setting and participants: For this retrospective study, we identified 4184 patients with unilateral surgically treated nonmetastatic pRCC in the Surveillance, Epidemiology, and End Results database (2004-2019). Outcome measurements and statistical analysis: The original VENUSS and GRANT risk categories were applied to predict 5-yr cancer-specific survival. A cross-validation method was used to test the accuracy and calibration of the models and to conduct decision curve analyses for the study cohort. Results and limitations: The VENUSS and GRANT categories represented independent predictors of cancer-specific mortality. On cross-validation, the accuracy of the VENUSS and GRANT risk categories was 0.73 and 0.65, respectively. Both models showed good calibration and performed better than random predictions in decision curve analysis. Limitations include the retrospective nature of the study and the absence of a central pathological review. Conclusion: VENUSS risk categories fulfilled prognostic model criteria for predicting cancer-specific survival 5 yr after surgery in North American patients with nonmetastatic pRCC as recommended by guidelines. Conversely, GRANT risk categories did not. Thus, VENUSS risk categories represent an important tool for counseling, follow-up planning, and patient selection for appropriate adjuvant trials in pRCC. Patient summary: We tested the ability of two validated methods (VENUSS and GRANT) to predict death due to papillary kidney cancer in a North American population. The VENUSS risk categories showed good performance in predicting 5-year cancer-specific survival.

Cancer-specific mortality free survival rates in non-metastatic non-clear cell renal carcinoma patients at intermediate/high risk of recurrence.

BACKGROUND: To date, five trials testing the effect of adjuvant systemic therapy in surgically treated non-metastatic renal cell carcinoma included patients with non-clear cell histology. We tested the effect of papillary vs. chromophobe histological subtype, stage, and grade on 10-year cancer-specific survival, in patients eligible for ≥1 such trial. METHODS: We identified patients meeting ASSURE, SORCE, EVEREST, PROSPER, or RAMPART trial inclusion criteria in the SEER (2000-2018) database. Kaplan-Meier analyses estimated 10-year survival rates and multivariable Cox regression models tested for the independent predictor status of histological subtype, stage, and grade. RESULTS: We identified 5465 (68%) papillary and 2562 (32%) chromophobe renal cell carcinoma patients. Cancer-specific survival rates at 10 years were 77% in papillary vs. 90% in chromophobe. In multivariable Cox regression models applied to papillary patients, cancer-specific mortality independent predictor status was reached for T3G3-4 (HR 2.9), T4Gany (HR 3.4), TanyN1G1-2 (HR 3.1), and TanyN1G3-4 (HR 8.0, P<0.001), relative to T1/2Gany. In multivariable Cox regression models applied to chromophobe patients, mortality independent predictor status was reached for T3G3-4 (HR 3.6), T4Gany (HR 14.0), TanyN1G1-2 (HR 5.7), and TanyN1G3-4 (HR 15.0, P<0.001), relative to T1/2Gany. CONCLUSIONS: In surgically treated non-metastatic intermediate/high-risk renal cell carcinoma patients, papillary histologic subtype exhibited worse cancer-specific survival than chromophobe histologic subtype. Although stage and grade represented independent predictors in both histological subtype groups, the magnitude of their effect was invariably worse in chromophobe than in papillary patients. In consequence, papillary and chromophobe patients should be considered separate entities instead of being combined under the non-clear cell designation.

Characterization of sinoatrial automaticity in Microcebus murinus to study the effect of aging on cardiac activity and the correlation with longevity.

Microcebus murinus, or gray mouse lemur (GML), is one of the smallest primates known, with a size in between mice and rats. The small size, genetic proximity to humans and prolonged senescence, make this lemur an emerging model for neurodegenerative diseases. For the same reasons, it could help understand how aging affects cardiac activity. Here, we provide the first characterization of sinoatrial (SAN) pacemaker activity and of the effect of aging on GML heart rate (HR). According to GML size, its heartbeat and intrinsic pacemaker frequencies lie in between those of mice and rats. To sustain this fast automaticity the GML SAN expresses funny and Ca2+ currents (If, ICa,L and ICa,T) at densities similar to that of small rodents. SAN automaticity was also responsive to ß-adrenergic and cholinergic pharmacological stimulation, showing a consequent shift in the localization of the origin of pacemaker activity. We found that aging causes decrease of basal HR and atrial remodeling in GML. We also estimated that, over 12 years of a lifetime, GML generates about 3 billion heartbeats, thus, as many as humans and three times more than rodents of equivalent size. In addition, we estimated that the high number of heartbeats per lifetime is a characteristic that distinguishes primates from rodents or other eutherian mammals, independently from body size. Thus, cardiac endurance could contribute to the exceptional longevity of GML and other primates, suggesting that GML's heart sustains a workload comparable to that of humans in a lifetime. In conclusion, despite the fast HR, GML replicates some of the cardiac deficiencies reported in old people, providing a suitable model to study heart rhythm impairment in aging. Moreover, we estimated that, along with humans and other primates, GML presents a remarkable cardiac longevity, enabling longer life span than other mammals of equivalent size.

Utility of PINP to monitor osteoporosis treatment in primary care, the POSE study (PINP and Osteoporosis in Sheffield Evaluation).

PURPOSE: In Sheffield (UK), we introduced the PINP monitoring algorithm for the management of osteoporosis treatment delivered in primary care. Our aims were to evaluate whether this algorithm was associated with better osteoporosis outcomes and was cost-effective compared to standard care. METHODS: Inclusion criteria were referral from Sheffield GPs, BMD scans performed between 2012 and 2013 and a report advising initiation of oral bisphosphonate and PINP monitoring. 906 patients were identified and retrospectively divided into Group A (intention to monitor, with baseline PINP, n = 588) and Group B (no intention to monitor, without baseline PINP, n = 318). The model described by Davis and colleagues was used to extrapolate life-time costs and quality-adjusted life-years (QALYs). RESULTS: No differences were found in baseline characteristics between groups (age, gender, BMI, BMD and major risk factors for fractures). More patients in Group A started oral treatment (77.4% vs 49.1%; p < 0.001), but there were no differences between groups in the presence of a gap in treatment >3 months or in treatment duration. Patients in Group A were more likely to have follow-up DXA scan at 4-6 years from baseline (46.9% vs 29.2%; p < 0.000) and had a greater increase in total hip BMD (+2.74% vs + 0.42%; p value = 0.003). Fewer new fractures occurred in Group A but this was not statistically significant, but the numbers of fractures were small. Patients in Group A were more likely to change management (p = 0.005) including switching to zoledronate (p = 0.03). The PINP measurement and increased prescribing in Group A resulted in increases in both costs (£30.19) and QALYs (0.0039) relative to Group B, giving an incremental cost effectiveness ratio (ICER) of £7660 in the probabilistic sensitivity analysis. CONCLUSIONS: Patients monitored with PINP are more likely to start oral bisphosphonate treatment, switch to zoledronate, have follow-up DXA scans and a greater increase of hip BMD. PINP monitoring has the potential to be cost-effective in a UK NHS setting given that interventions with an ICER under £20,000 are generally considered to be cost-effective.

The funny current in genetically modified mice.

Since its first description in 1979, the hyperpolarization-activated funny current (If) has been the object of intensive research aimed at understanding its role in cardiac pacemaker activity and its modulation by the sympathetic and parasympathetic branches of the autonomic nervous system. If was described in isolated tissue strips of the rabbit sinoatrial node using the double-electrode voltage-clamp technique. Since then, the rabbit has been the principal animal model for studying pacemaker activity and If for more than 20 years. In 2001, the first study describing the electrophysiological properties of mouse sinoatrial pacemaker myocytes and those of If was published. It was soon followed by the description of murine myocytes of the atrioventricular node and the Purkinje fibres. The sinoatrial node of genetically modified mice has become a very popular model for studying the mechanisms of cardiac pacemaker activity. This field of research benefits from the impressive advancement of in-vivo exploration techniques of physiological parameters, imaging, genetics, and large-scale genomic approaches. The present review discusses the influence of mouse genetic on the most recent knowledge of the funny current's role in the physiology and pathophysiology of cardiac pacemaker activity. Genetically modified mice have provided important insights into the role of If in determining intrinsic automaticity in vivo and in myocytes of the conduction system. In addition, gene targeting of f-(HCN) channel isoforms have contributed to elucidating the current's role in the regulation of heart rate by the parasympathetic nervous system. This review is dedicated to Dario DiFrancesco on his retirement.

Novel roles for voltage-gated T-type Ca2+ and ClC-2 channels in phagocytosis and angiogenic factor balance identified in human iPSC-derived RPE.

Human-induced pluripotent stem cell (hiPSC)-derived retinal pigment epithelium (RPE) is a powerful tool for pathophysiological studies and preclinical therapeutic screening, as well as a source for clinical cell transplantation. Thus, it must be validated for maturity and functionality to ensure correct data readouts and clinical safety. Previous studies have validated hiPSC-derived RPE as morphologically characteristic of the tissue in the human eye. However, information concerning the expression and functionality of ion channels is still limited. We screened hiPSC-derived RPE for the polarized expression of a panel of L-type (CaV 1.1, CaV 1.3) and T-type (CaV 3.1, CaV 3.3) Ca2+ channels, K+ channels (Maxi-K, Kir4.1, Kir7.1), and the Cl- channel ClC-2 known to be expressed in native RPE. We also tested the roles of these channels in key RPE functions using specific inhibitors. In addition to confirming the native expression profiles and function of certain channels, such as L-type Ca2+ channels, we show for the first time that T-type Ca2+ channels play a role in both phagocytosis and vascular endothelial growth factor (VEGF) secretion. Moreover, we demonstrate that Maxi-K and Kir7.1 channels are involved in the polarized secretion of VEGF and pigment epithelium-derived factor (PEDF). Furthermore, we show a novel localization for ClC-2 channel on the apical side of hiPSC-derived RPE, with an overexpression at the level of fluid-filled domes, and demonstrate that it plays an important role in phagocytosis, as well as VEGF and PEDF secretion. Taken together, hiPSC-derived RPE is a powerful model for advancing fundamental knowledge of RPE functions.

Subretinally injected semiconducting polymer nanoparticles rescue vision in a rat model of retinal dystrophy.

Inherited retinal dystrophies and late-stage age-related macular degeneration, for which treatments remain limited, are among the most prevalent causes of legal blindness. Retinal prostheses have been developed to stimulate the inner retinal network; however, lack of sensitivity and resolution, and the need for wiring or external cameras, have limited their application. Here we show that conjugated polymer nanoparticles (P3HT NPs) mediate light-evoked stimulation of retinal neurons and persistently rescue visual functions when subretinally injected in a rat model of retinitis pigmentosa. P3HT NPs spread out over the entire subretinal space and promote light-dependent activation of spared inner retinal neurons, recovering subcortical, cortical and behavioural visual responses in the absence of trophic effects or retinal inflammation. By conferring sustained light sensitivity to degenerate retinas after a single injection, and with the potential for high spatial resolution, P3HT NPs provide a new avenue in retinal prosthetics with potential applications not only in retinitis pigmentosa, but also in age-related macular degeneration.

Diamond-inclusion system recording old deep lithosphere conditions at Udachnaya (Siberia).

Diamonds and their inclusions are unique fragments of deep Earth, which provide rare samples from inaccessible portions of our planet. Inclusion-free diamonds cannot provide information on depth of formation, which could be crucial to understand how the carbon cycle operated in the past. Inclusions in diamonds, which remain uncorrupted over geological times, may instead provide direct records of deep Earth's evolution. Here, we applied elastic geothermobarometry to a diamond-magnesiochromite (mchr) host-inclusion pair from the Udachnaya kimberlite (Siberia, Russia), one of the most important sources of natural diamonds. By combining X-ray diffraction and Fourier-transform infrared spectroscopy data with a new elastic model, we obtained entrapment conditions, Ptrap = 6.5(2) GPa and Ttrap = 1125(32)-1140(33) °C, for the mchr inclusion. These conditions fall on a ca. 35 mW/m2 geotherm and are colder than the great majority of mantle xenoliths from similar depth in the same kimberlite. Our results indicate that cold cratonic conditions persisted for billions of years to at least 200 km in the local lithosphere. The composition of the mchr also indicates that at this depth the lithosphere was, at least locally, ultra-depleted at the time of diamond formation, as opposed to the melt-metasomatized, enriched composition of most xenoliths.

Performance and clinical comparison between left ventricular quadripolar and bipolar leads in cardiac resynchronization therapy: Observational research.

AIM: To evaluate Attain Performa (Medtronic, Dublin, Ireland) quadripolar lead performance in clinical practice and, secondarily, to compare its long term clinical outcomes vs bipolar leads for left ventricular (LV) pacing. METHODS AND RESULTS: We retrospectively analyzed clinical, procedural and follow-up data of 215 patients implanted with a quadripolar lead. One hundred and twenty one patients implanted with bipolar lead were selected to compare long-term clinical outcomes. The quadripolar lead was implanted in the target vein in 196 patients (91%) without acute dislodgements. In 50% of patients the chosen final pacing configuration at implant would not have been available with bipolar leads. A dedicated quadripolar pacing vector was chosen more frequently when the LV tip location was apical than otherwise (65.6% vs 42.7%, p=0.003). After a median follow-up of 14 months, the LV pacing threshold was less than 2.5V at 0.4ms in 98 patients (90%) with a safety margin between phrenic nerve and LV pacing threshold >3V in 97 patients (89%). We observed a slight trend toward a lower risk of heart failure worsening and a lower incidence of ventricular arrhythmias and pulmonary congestion in patients implanted with quadripolar leads compared with the control group. CONCLUSION: Quadripolar leads improve the management of phrenic nerve stimulation at no trade-off with pacing threshold and lead stability. Quadripolar leads seems to be associated with a lower incidence of VT/VF and pulmonary congestion, when compared with bipolar leads, but further investigations are necessary to confirm that this positive effect is associated with better LV reverse remodeling.

Interfacing Graphene-Based Materials With Neural Cells.

The scientific community has witnessed an exponential increase in the applications of graphene and graphene-based materials in a wide range of fields, from engineering to electronics to biotechnologies and biomedical applications. For what concerns neuroscience, the interest raised by these materials is two-fold. On one side, nanosheets made of graphene or graphene derivatives (graphene oxide, or its reduced form) can be used as carriers for drug delivery. Here, an important aspect is to evaluate their toxicity, which strongly depends on flake composition, chemical functionalization and dimensions. On the other side, graphene can be exploited as a substrate for tissue engineering. In this case, conductivity is probably the most relevant amongst the various properties of the different graphene materials, as it may allow to instruct and interrogate neural networks, as well as to drive neural growth and differentiation, which holds a great potential in regenerative medicine. In this review, we try to give a comprehensive view of the accomplishments and new challenges of the field, as well as which in our view are the most exciting directions to take in the immediate future. These include the need to engineer multifunctional nanoparticles (NPs) able to cross the blood-brain-barrier to reach neural cells, and to achieve on-demand delivery of specific drugs. We describe the state-of-the-art in the use of graphene materials to engineer three-dimensional scaffolds to drive neuronal growth and regeneration in vivo, and the possibility of using graphene as a component of hybrid composites/multi-layer organic electronics devices. Last but not least, we address the need of an accurate theoretical modeling of the interface between graphene and biological material, by modeling the interaction of graphene with proteins and cell membranes at the nanoscale, and describing the physical mechanism(s) of charge transfer by which the various graphene materials can influence the excitability and physiology of neural cells.

Effect of cytosolic pH on inward currents reveals structural characteristics of the proton transport cycle in the influenza A protein M2 in cell-free membrane patches of Xenopus oocytes.

Transport activity through the mutant D44A of the M2 proton channel from influenza virus A was measured in excised inside-out macro-patches of Xenopus laevis oocytes at cytosolic pH values of 5.5, 7.5 and 8.2. The current-voltage relationships reveal some peculiarities: 1. "Transinhibition", i.e., instead of an increase of unidirectional outward current with increasing cytosolic H(+) concentration, a decrease of unidirectional inward current was found. 2. Strong inward rectification. 3. Exponential rise of current with negative potentials. In order to interpret these findings in molecular terms, different kinetic models have been tested. The transinhibition basically results from a strong binding of H(+) to a site in the pore, presumably His37. This assumption alone already provides inward rectification and exponential rise of the IV curves. However, it results in poor global fits of the IV curves, i.e., good fits were only obtained for cytosolic pH of 8.2, but not for 7.5. Assuming an additional transport step as e.g. caused by a constriction zone at Val27 resulted in a negligible improvement. In contrast, good global fits for cytosolic pH of 7.5 and 8.2 were immediately obtained with a cyclic model. A "recycling step" implies that the protein undergoes conformational changes (assigned to Trp41 and Val27) during transport which have to be reset before the next proton can be transported. The global fit failed at the low currents at pHcyt = 5.5, as expected from the interference of putative transport of other ions besides H(+). Alternatively, a regulatory effect of acidic cytosolic pH may be assumed which strongly modifies the rate constants of the transport cycle.

The G-protein-gated K+ channel, IKACh, is required for regulation of pacemaker activity and recovery of resting heart rate after sympathetic stimulation.

Parasympathetic regulation of sinoatrial node (SAN) pacemaker activity modulates multiple ion channels to temper heart rate. The functional role of the G-protein-activated K(+) current (IKACh) in the control of SAN pacemaking and heart rate is not completely understood. We have investigated the functional consequences of loss of IKACh in cholinergic regulation of pacemaker activity of SAN cells and in heart rate control under physiological situations mimicking the fight or flight response. We used knockout mice with loss of function of the Girk4 (Kir3.4) gene (Girk4(-/-) mice), which codes for an integral subunit of the cardiac IKACh channel. SAN pacemaker cells from Girk4(-/-) mice completely lacked IKACh. Loss of IKACh strongly reduced cholinergic regulation of pacemaker activity of SAN cells and isolated intact hearts. Telemetric recordings of electrocardiograms of freely moving mice showed that heart rate measured over a 24-h recording period was moderately increased (10%) in Girk4(-/-) animals. Although the relative extent of heart rate regulation of Girk4(-/-) mice was similar to that of wild-type animals, recovery of resting heart rate after stress, physical exercise, or pharmacological ß-adrenergic stimulation of SAN pacemaking was significantly delayed in Girk4(-/-) animals. We conclude that IKACh plays a critical role in the kinetics of heart rate recovery to resting levels after sympathetic stimulation or after direct ß-adrenergic stimulation of pacemaker activity. Our study thus uncovers a novel role for IKACh in SAN physiology and heart rate regulation.

The proapoptotic influenza A virus protein PB1-F2 forms a nonselective ion channel.

BACKGROUND: PB1-F2 is a proapoptotic influenza A virus protein of approximately 90 amino acids in length that is located in the nucleus, cytosol and in the mitochondria membrane of infected cells. Previous studies indicated that the molecule destabilizes planar lipid bilayers and has a strong inherent tendency for multimerization. This may be correlate with its capacity to induce mitochondrial membrane depolarization. METHODOLOGY/PRINCIPAL FINDINGS: Here, we investigated whether PB1-F2 is able to form ion channels within planar lipid bilayers and microsomes. For that purpose, a set of biologically active synthetic versions of PB1-F2 (sPB1-F2) derived from the IAV isolates A/Puerto Rico/8/34(H1N1) (IAV(PR8)), from A/Brevig Mission/1/1918(H1N1) (IAV(SF2)) or the H5N1 consensus sequence (IAV(BF2)) were used. Electrical and fluorimetric measurements show that all three peptides generate in planar lipid bilayers or in liposomes, respectively, a barely selective conductance that is associated with stochastic channel type fluctuations between a closed state and at least two defined open states. Unitary channel fluctuations were also generated when a truncated protein comprising only the 37 c-terminal amino acids of sPB1-F2 was reconstituted in bilayers. Experiments were complemented by extensive molecular dynamics simulations of the truncated fragment in a lipid bilayer. The results indicate that the c-terminal region exhibits a slightly bent helical fold, which is stable and remains embedded in the bilayer for over 180 ns. CONCLUSION/SIGNIFICANCE: The data support the idea that PB1-F2 is able to form protein channel pores with no appreciable selectivity in membranes and that the c-terminus is important for this function. This information could be important for drug development.