Cellular migration into a subretinal honeycomb-shaped prosthesis for high-resolution prosthetic vision.

In patients blinded by geographic atrophy, a subretinal photovoltaic implant with 100 µm pixels provided visual acuity closely matching the pixel pitch. However, such flat bipolar pixels cannot be scaled below 75 µm, limiting the attainable visual acuity. This limitation can be overcome by shaping the electric field with 3-dimensional (3-D) electrodes. In particular, elevating the return electrode on top of the honeycomb-shaped vertical walls surrounding each pixel extends the electric field vertically and decouples its penetration into tissue from the pixel width. This approach relies on migration of the retinal cells into the honeycomb wells. Here, we demonstrate that majority of the inner retinal neurons migrate into the 25 µm deep wells, leaving the third-order neurons, such as amacrine and ganglion cells, outside. This enables selective stimulation of the second-order neurons inside the wells, thus preserving the intraretinal signal processing in prosthetic vision. Comparable glial response to that with flat implants suggests that migration and separation of the retinal cells by the walls does not cause additional stress. Furthermore, retinal migration into the honeycombs does not negatively affect its electrical excitability, while grating acuity matches the pixel pitch down to 40 µm and reaches the 27 µm limit of natural resolution in rats with 20 µm pixels. These findings pave the way for 3-D subretinal prostheses with pixel sizes of cellular dimensions.

[Age-dependent expression of HSP90 in the hippocampus of APP/PS1 mice].

The present study aims to observe the change in expression of heat shock protein 90 (HSP90) along with amyloid-ß (Aß) and phosphorylated Tau (p-Tau) protein levels in the hippocampus tissue of Alzheimer's disease (AD) transgenic animal model with age. APP/PS1 transgenic mice at age of 6-, 9- and 12-month and C57BL/6J mice of the same age were used. The cognitive abilities of these animals were evaluated using a Morris water maze. Western blot or immunohistochemistry was used to detect the expressions of HSP90 and Aß1-42, as well as the phosphorylation levels of Tau protein in the hippocampus. The hsp90 mRNA levels and the morphology and number of cells in the hippocampus were detected with real-time quantitative polymerase chain reaction (qRT-PCR) and Nissl staining, respectively. The results showed that compared with C57BL/6J mice of the same age, HSP90 and hsp90 mRNA expression were decreased (P < 0.05 or P < 0.01), while Aß1-42 and p-Tau protein levels were increased (P < 0.05 or P < 0.01) in the hippocampal tissue of APP/PS1 transgenic mice. Meanwhile, the decrease in HSP90 and hsp90 mRNA expression (P < 0.05 or P < 0.01), the increase in Aß1-42 and p-Tau levels (P < 0.01 or P < 0.05) in hippocampal tissue and the reduction in behavioral ability showed a progressive development with the advancing of age in the APP/PS1 transgenic mice. In conclusion, in the hippocampal tissue of APP/PS1 mice, the decrease in HSP90 expression and the increase in Aß1-42 and p-Tau levels together with the decline of their cognitive ability are age-dependent.

Three-dimensional electro-neural interfaces electroplated on subretinal prostheses.

Objective.Retinal prosthetics offer partial restoration of sight to patients blinded by retinal degenerative diseases through electrical stimulation of the remaining neurons. Decreasing the pixel size enables increasing prosthetic visual acuity, as demonstrated in animal models of retinal degeneration. However, scaling down the size of planar pixels is limited by the reduced penetration depth of the electric field in tissue. We investigated 3-dimensional (3d) structures on top of photovoltaic arrays for enhanced penetration of the electric field, permitting higher resolution implants.Approach.3D COMSOL models of subretinal photovoltaic arrays were developed to accurately quantify the electrodynamics during stimulation and verified through comparison to flat photovoltaic arrays. Models were applied to optimize the design of 3D electrode structures (pillars and honeycombs). Return electrodes on honeycomb walls vertically align the electric field with bipolar cells for optimal stimulation. Pillars elevate the active electrode, thus improving proximity to target neurons. The optimized 3D structures were electroplated onto existing flat subretinal prostheses.Main results.Simulations demonstrate that despite exposed conductive sidewalls, charge mostly flows via high-capacitance sputtered iridium oxide films topping the 3D structures. The 24µm height of honeycomb structures was optimized for integration with the inner nuclear layer cells in the rat retina, whilst 35µm tall pillars were optimized for penetrating the debris layer in human patients. Implantation of released 3D arrays demonstrates mechanical robustness, with histology demonstrating successful integration of 3D structures with the rat retinain-vivo.Significance. Electroplated 3D honeycomb structures produce vertically oriented electric fields, providing low stimulation thresholds, high spatial resolution, and high contrast for pixel sizes down to 20µm. Pillar electrodes offer an alternative for extending past the debris layer. Electroplating of 3D structures is compatible with the fabrication process of flat photovoltaic arrays, enabling much more efficient retinal stimulation.

3D electronic implants in subretinal space: Long-term follow-up in rodents.

Clinical results with photovoltaic subretinal prosthesis (PRIMA) demonstrated restoration of sight via electrical stimulation of the interneurons in degenerated retina, with resolution matching the 100 µm pixel size. Since scaling the pixels below 75 µm in the current bipolar planar geometry will significantly limit the penetration depth of the electric field and increase stimulation threshold, we explore the possibility of using smaller pixels based on a novel 3-dimensional honeycomb-shaped design. We assessed the long-term biocompatibility and stability of these arrays in rats by investigating the anatomical integration of the retina with flat and 3D implants and response to electrical stimulation over lifetime - up to 32-36 weeks post-implantation in aged rats. With both flat and 3D implants, signals elicited in the visual cortex decreased after the day of implantation by more than 3-fold, and gradually recovered over the next 12-16 weeks. With 25 µm high honeycomb walls, the majority of bipolar cells migrate into the wells, while amacrine and ganglion cells remain above the cavities, which is essential for selective network-mediated stimulation of the retina. Retinal thickness and full-field stimulation threshold with 40 µm-wide honeycomb pixels were comparable to those with planar devices - 0.05 mW/mm2 with 10 ms pulses. However, fewer cells from the inner nuclear layer migrated into the 20 µm-wide wells, and stimulation threshold increased over 12-16 weeks, before stabilizing at about 0.08 mW/mm2. Such threshold is still significantly lower than 1.8 mW/mm2 with a previous design of flat bipolar pixels, confirming the promise of the 3D honeycomb-based approach to high resolution subretinal prosthesis.

Three-dimensional electro-neural interfaces electroplated on subretinal prostheses.

Objective: High-resolution retinal prosthetics offer partial restoration of sight to patients blinded by retinal degenerative diseases through electrical stimulation of the remaining neurons. Decreasing the pixel size enables an increase in prosthetic visual acuity, as demonstrated in animal models of retinal degeneration. However, scaling down the size of planar pixels is limited by the reduced penetration depth of the electric field in tissue. We investigate 3-dimensional structures on top of the photovoltaic arrays for enhanced penetration of electric field to permit higher-resolution implants. Approach: We developed 3D COMSOL models of subretinal photovoltaic arrays that accurately quantify the device electrodynamics during stimulation and verified it experimentally through comparison with the standard (flat) photovoltaic arrays. The models were then applied to optimise the design of 3D electrode structures (pillars and honeycombs) to efficiently stimulate the inner retinal neurons. The return electrodes elevated on top of the honeycomb walls surrounding each pixel orient the electric field inside the cavities vertically, aligning it with bipolar cells for optimal stimulation. Alternatively, pillars elevate the active electrode into the inner nuclear layer, improving proximity to the target neurons. Modelling results informed a microfabrication process of electroplating the 3D electrode structures on top of the existing flat subretinal prosthesis. Main results: Simulations demonstrate that despite the conductive sidewalls of the 3D electrodes being exposed to electrolyte, most of the charge flows via the high-capacitance sputtered Iridium Oxide film that caps the top of the 3D structures. The 24 µm height of the electroplated honeycomb structures was optimised for integration with the inner nuclear layer cells in rat retina, while 35 µm height of the pillars was optimized for penetrating the debris layer in human patients. Release from the wafer and implantation of the 3D arrays demonstrated that they are mechanically robust to withstand the associated forces. Histology demonstrated successful integration of the 3D structures with the rat retina in-vivo. Significance: Electroplated 3D honeycomb structures produce a vertically oriented electric field that offers low stimulation threshold, high spatial resolution and high contrast for the retinal implants with pixel sizes down to 20µm in width. Pillar electrodes offer an alternative configuration for extending the stimulation past the debris layers. Electroplating of the 3D structures is compatible with the fabrication process of the flat photovoltaic arrays, thereby enabling much more efficient stimulation than in their original flat configuration.

3D electronic implants in subretinal space: long-term follow-up in rodents.

Photovoltaic subretinal prosthesis (PRIMA) enables restoration of sight via electrical stimulation of the interneurons in degenerated retina, with resolution limited by the 100 µm pixel size. Since decreasing the pixel size below 75 µm in the current bipolar geometry is impossible, we explore the possibility of using smaller pixels based on a novel 3-dimensional honeycomb-shaped design. We assessed the long-term biocompatibility and stability of these arrays in rats by investigating the anatomical integration of the retina with flat and 3D implants and response to electrical stimulation over lifetime - up to 9 months post-implantation in aged rats. With both flat and 3D implants, VEP amplitude decreased after the day of implantation by more than 3-fold, and gradually recovered over about 3 months. With 25 µm high honeycomb walls, the majority of bipolar cells migrate into the wells, while amacrine and ganglion cells remain above the cavities, which is essential for selective network-mediated stimulation of the second-order neurons. Retinal thickness and full-field stimulation threshold with 40 µm-wide honeycomb pixels were comparable to those with planar devices - 0.05 mW/mm2 with 10ms pulses. However, fewer cells from the inner nuclear layer migrated into the 20 µm-wide wells, and stimulation threshold increased over 5 months, before stabilizing at about 0.08 mW/mm2. Such threshold is significantly lower than 1.8 mW/mm2 with a previous design of flat bipolar pixels, confirming the promise of the 3D honeycomb-based approach to high resolution subretinal prosthesis.

Pixel size limit of the PRIMA implants: from humans to rodents and back.

Objective.Retinal prostheses aim at restoring sight in patients with retinal degeneration by electrically stimulating the inner retinal neurons. Clinical trials with patients blinded by atrophic age-related macular degeneration using the PRIMA subretinal implant, a 2 × 2 mm array of 100µm-wide photovoltaic pixels, have demonstrated a prosthetic visual acuity closely matching the pixel size. Further improvement in resolution requires smaller pixels, which, with the current bipolar design, necessitates more intense stimulation.Approach.We examine the lower limit of the pixel size for PRIMA implants by modeling the electric field, leveraging the clinical benchmarks, and using animal data to assess the stimulation strength and contrast of various patterns. Visually evoked potentials measured in Royal College of Surgeons rats with photovoltaic implants composed of 100µm and 75µm pixels were compared to clinical thresholds with 100µm pixels. Electrical stimulation model calibrated by the clinical and rodent data was used to predict the performance of the implant with smaller pixels.Main results.PRIMA implants with 75µm bipolar pixels under the maximum safe near-infrared (880 nm) illumination of 8 mW mm-2with 30% duty cycle (10 ms pulses at 30 Hz) should provide a similar perceptual brightness as with 100µm pixels under 3 mW mm-2irradiance, used in the current clinical trials. Contrast of the Landolt C pattern scaled down to 75µm pixels is also similar under such illumination to that with 100µm pixels, increasing the maximum acuity from 20/420 to 20/315.Significance.Computational modeling defines the minimum pixel size of the PRIMA implants as 75µm. Increasing the implant width from 2 to 3 mm and reducing the pixel size from 100 to 75µm will nearly quadrupole the number of pixels, which should be very beneficial for patients. Smaller pixels of the same bipolar flat geometry would require excessively intense illumination, and therefore a different pixel design should be considered for further improvement in resolution.

Photovoltaic implant simulator reveals resolution limits in subretinal prosthesis.

Objective.PRIMA, the photovoltaic subretinal prosthesis, restores central vision in patients blinded by atrophic age-related macular degeneration (AMD), with a resolution closely matching the 100µm pixel size of the implant. Improvement in resolution requires smaller pixels, but the resultant electric field may not provide sufficient stimulation strength in the inner nuclear layer (INL) or may lead to excessive crosstalk between neighboring electrodes, resulting in low contrast stimulation patterns. We study the approaches to electric field shaping in the retina for prosthetic vision with higher resolution and improved contrast.Approach.We present a new computational framework, Retinal Prosthesis Simulator (RPSim), that efficiently computes the electric field in the retina generated by a photovoltaic implant with thousands of electrodes. Leveraging the PRIMA clinical results as a benchmark, we use RPSim to predict the stimulus strength and contrast of the electric field in the retina with various pixel designs and stimulation patterns.Main results.We demonstrate that by utilizing monopolar pixels as both anodes and cathodes to suppress crosstalk, most patients may achieve resolution no worse than 48µm. Closer proximity between the electrodes and the INL, achieved with pillar electrodes, enhances the stimulus strength and contrast and may enable 24µm resolution with 20µm pixels, at least in some patients.Significance.A resolution of 24µm on the retina corresponds to a visual acuity of 20/100, which is over 4 times higher than the current best prosthetic acuity of 20/438, promising a significant improvement of central vision for many AMD patients.

Electronic photoreceptors enable prosthetic visual acuity matching the natural resolution in rats.

Localized stimulation of the inner retinal neurons for high-acuity prosthetic vision requires small pixels and minimal crosstalk from the neighboring electrodes. Local return electrodes within each pixel limit the crosstalk, but they over-constrain the electric field, thus precluding the efficient stimulation with subretinal pixels smaller than 55 µm. Here we demonstrate a high-resolution prosthetic vision based on a novel design of a photovoltaic array, where field confinement is achieved dynamically, leveraging the adjustable conductivity of the diodes under forward bias to turn the designated pixels into transient returns. We validated the computational modeling of the field confinement in such an optically-controlled circuit by in-vitro and in-vivo measurements. Most importantly, using this strategy, we demonstrated that the grating acuity with 40 µm pixels matches the pixel pitch, while with 20 µm pixels, it reaches the 28 µm limit of the natural visual resolution in rats. This method enables customized field shaping based on individual retinal thickness and distance from the implant, paving the way to higher acuity of prosthetic vision in atrophic macular degeneration.

Vertical-junction photodiodes for smaller pixels in retinal prostheses.

Objective.To restore central vision in patients with atrophic age-related macular degeneration, we replace the lost photoreceptors with photovoltaic pixels, which convert light into current and stimulate the secondary retinal neurons. Clinical trials demonstrated prosthetic acuity closely matching the sampling limit of the 100µm pixels, and hence smaller pixels are required for improving visual acuity. However, with smaller flat bipolar pixels, the electric field penetration depth and the photodiode responsivity significantly decrease, making the device inefficient. Smaller pixels may be enabled by (a) increasing the diode responsivity using vertical p-n junctions and (b) directing the electric field in tissue vertically. Here, we demonstrate such novel photodiodes and test the retinal stimulation in a vertical electric field.Approach.Arrays of silicon photodiodes of 55, 40, 30, and 20µm in width, with vertical p-n junctions, were fabricated. The electric field in the retina was directed vertically using a common return electrode at the edge of the device. Optical and electronic performance of the diodes was characterizedin-vitro, and retinal stimulation threshold measured by recording the visually evoked potentials in rats with retinal degeneration.Main results.The photodiodes exhibited sufficiently low dark current (<10 pA) and responsivity at 880 nm wavelength as high as 0.51 A W-1, with 85% internal quantum efficiency, independent of pixel size. Field mapping in saline demonstrated uniformity of the pixel performance in the array. The full-field stimulation threshold was as low as 0.057±0.029mW mm-2with 10 ms pulses, independent of pixel size.Significance.Photodiodes with vertical p-n junctions demonstrated excellent charge collection efficiency independent of pixel size, down to 20µm. Vertically oriented electric field provides a stimulation threshold that is independent of pixel size. These results are the first steps in validation of scaling down the photovoltaic pixels for subretinal stimulation.

Harmonic-balance circuit analysis for electro-neural interfaces.

OBJECTIVE: Avoidance of the adverse electrochemical reactions at the electrode-electrolyte interface defines the voltage safety window and limits the charge injection capacity (CIC) of an electrode material. For an electrode that is not ideally capacitive, the CIC depends on the waveform of the stimulus. We study the modeling of the charge injection dynamics to optimize the waveforms for efficient neural stimulation within the electrochemical safety limits. APPROACH: The charge injection dynamics at the electrode-electrolyte interface is typically characterized by the electrochemical impedance spectrum, and is often approximated by discrete-element circuit models. We compare the modeling of the complete circuit, including a non-linear driver such as a photodiode, based on the harmonic-balance (HB) analysis with the analysis based on various - (discrete-element) approximations. To validate the modeling results, we performed experiments with iridium-oxide electrodes driven by a current source with diodes in parallel, which mimics a photovoltaic circuit. MAIN RESULTS: Application of HB analysis based on a full impedance spectrum eliminates the complication of finding the discrete-element circuit model in traditional approaches. HB-based results agree with the experimental data better than the discrete-element circuit. HB technique can be applied not only to demonstrate the circuit response to periodic stimulation, but also to describe the initial transient behavior when a burst waveform is applied. SIGNIFICANCE: HB-based circuit analysis accurately describes the dynamics of electrode-electrolyte interfaces and driving circuits for all pulsing schemes. This allows optimizing the stimulus waveform to maximize the CIC, based on the impedance spectrum alone.

On optimal coupling of the 'electronic photoreceptors' into the degenerate retina.

Objective: To restore sight in atrophic age-related macular degeneration, the lost photoreceptors can be replaced with electronic implants, which replicate their two major functions: (1) converting light into an electric signal, and (2) transferring visual information to the secondary neurons in the retinal neural network­the bipolar cells (BC). We study the selectivity of BC activation by subretinal implants and dynamics of their response to pulsatile waveforms in order to optimize the electrical stimulation scheme such that retinal signal processing with 'electronic photoreceptors' remains as close to natural as possible. Approach: A multicompartmental model of a BC was implemented to simulate responses of the voltage-gated calcium channels and subsequent synaptic vesicle release under continuous and pulsatile stimuli. We compared the predicted response under various frequencies, pulse durations, and alternating gratings to the corresponding experimental measurements. In addition, electric field was computed for various electrode configurations in a 3-d finite element model to assess the stimulation selectivity via spatial confinement of the field. Main results: The modeled BC-mediated retinal responses were, in general, in good agreement with previously published experimental results. Kinetics of the calcium pumps and of the neurotransmitter release in ribbon synapses, which underpin the BC's temporal filtering and rectifying functions, allow mimicking the natural BC response with high frequency pulsatile stimulation, thereby preserving features of the retinal signal processing, such as flicker fusion, adaptation to static stimuli and non-linear summation of subunits in receptive field. Selectivity of the BC stimulation while avoiding direct activation of the downstream neurons (amacrine and ganglion cells­RGCs) is improved with local return electrodes. Significance: If the retinal neural network is preserved to a large extent in age-related macular degeneration, selective stimulation of BCs with proper spatial and temporal modulation of the extracellular electric field may retain many features of the natural retinal signal processing and hence allow highly functional restoration of sight.

Effect of Papain on the Demulsification of Peanut Oil Body Emulsion and the Corresponding Mechanism.

This study investigated the effect of papain on the demulsification of peanut oil body emulsion extracted using an aqueous enzymatic method and the associated mechanism. The highest free oil yield using papain (92.39%) was obtained under the following conditions: an enzymatic hydrolysis temperature of 55°C, sample-to-water ratio of 1:3, enzyme concentration of 1400 U/g, and an enzymatic hydrolysis time of 3 h. Papain degraded the peanut oil body protein to small-molecular-weight peptides (≤ 14.4 kDa). Compared to the emulsion before enzymatic hydrolysis, the amino acid content in the aqueous phase was higher after enzymatic hydrolysis, the viscosity of the oil body emulsion was lower, and the particle diameter of the emulsion was significantly larger. The following demulsification mechanism was derived. Papain degrades the protein on the peanut oil body and dissolves it in water. The outer side of the oil body loses the protection of electrostatic repulsion and steric hindrance provided by the membrane protein. This causes the viscosity of the emulsion system and the molecular steric hindrance to decrease. As a result, the oil droplets gather and eventually demulsify. The results of this study provide the theoretical basis for the instability in oil body emulsions and are expected to promote the application of enzymatic demulsification in industry.

Dynamical Strain-Driven Phase Separation in Flexible CoFe2O4/CoO Exchange Coupling System.

Epitaxial CoFe2O4(CFO)/CoO bilayers were fabricated by pulsed laser deposition on flexible muscovite mica substrate. Samples with different CFO thicknesses were employed to study the phenomenon of exchange bias involving strongly anisotropic ferromagnet. Magnetic measurements exhibited great enhancement in the features of exchange bias. Raman and X-ray absorption spectroscopies indicated that a new phase emerged within the CFO layer because of the cation charge redistribution in CFO layer under bending, which in turn gave rise to anomalous hysteresis loops exhibited in the bent bilayers. These results provide a fundamental understanding about the mechanisms of exchange bias prevailing in these bilayers and call attention to the implementation of spintronic devices using flexible heterostructures such as the present CFO/CoO bilayers.

Development and implementation of a dynamically updated big data intelligence platform from electronic health records for nasopharyngeal carcinoma research.

OBJECTIVE: To develop a big data intelligence platform for secondary use of electronic health records (EHRs) data to facilitate research for nasopharyngeal cancer (NPC). METHODS: This project was launched in 2015 and carried out by the cooperation of an academic cancer centre and a technology company. Patients diagnosed with NPC at Sun Yat-sen University Cancer Centre since January 2008 were included in the platform. Standard data elements were established to defined 981 variables for the platform. For each patient, data from 13 EHRs systems were extracted, integrated, structurized and normalized. Eight functional modules were constructed for the platform to facilitate the investigators to identify eligible patients, establish research projects, conduct statistical analysis, track the follow-up, search literature, etc. RESULTS: From January 2008 to December 2018, 54,703 patients diagnosed with NPC were included. Of these patients, 39,058 (71.4%) were male, and 15,645 (28.6%) were female; median age was 47 (interquartile range, 39-55) years. Of 981 variables, 341 were obtained from data structurization and normalization, of which 68 were generated by interacting multiple data sources via well-defined logical rules. The average precision rate, recall rate and F-measure for 341 variables were 0.97 ± 0.024, 0.92 ± 0.030, and 0.94 ± 0.027 respectively. The platform is regularly updated every seven days to include new patients and add new data for existing patients. Up to now, eight big data-driven retrospective studies have been published from the platform. CONCLUSION: Our big data intelligence platform demonstrates the feasibility of integrating EHRs data of routine healthcare, and offers an important perspective on real-world study of NPC. The continued efforts may be focus on data sharing among multiple hospitals and publicly releasing of data files. ADVANCES IN KNOWLEDGE: Our big data intelligence platform is the first disease-specific data platform for NPC research. It incorporates comprehensive EHRs data from routine healthcare, which can facilitate real-world study of NPC in risk stratification, decision-making and comorbidities management.

Big-data analysis: A clinical pathway on endoscopic retrograde cholangiopancreatography for common bile duct stones.

BACKGROUND: A clinical pathway (CP) is a standardized approach for disease management. However, big data-based evidence is rarely involved in CP for related common bile duct (CBD) stones, let alone outcome comparisons before and after CP implementation. AIM: To investigate the value of CP implementation in patients with CBD stones undergoing endoscopic retrograde cholangiopancreatography (ERCP). METHODS: This retrospective study was conducted at Nanjing Drum Tower Hospital in patients with CBD stones undergoing ERCP from January 2007 to December 2017. The data and outcomes were compared by using univariate and multivariable regression/linear models between the patients who received conventional care (non-pathway group, n = 467) and CP care (pathway group, n = 2196). RESULTS: At baseline, the main differences observed between the two groups were the percentage of patients with multiple stones (P < 0.001) and incidence of cholangitis complication (P < 0.05). The percentage of antibiotic use and complications in the CP group were significantly less than those in the non-pathway group [adjusted odds ratio (OR) = 0.72, 95% confidence interval (CI): 0.55-0.93, P = 0.012, adjusted OR = 0.44, 95%CI: 0.33-0.59, P < 0.001, respectively]. Patients spent lower costs on hospitalization, operation, nursing, medication, and medical consumable materials (P < 0.001 for all), and even experienced shorter length of hospital stay (LOHS) (P < 0.001) after the CP implementation. No significant differences in clinical outcomes, readmission rate, or secondary surgery rate were presented between the patients in the non-pathway and CP groups. CONCLUSION: Implementing a CP for patients with CBD stones is a safe mode to reduce the LOHS, hospital costs, antibiotic use, and complication rate.

Transcriptome analysis of maca (Lepidium meyenii) root at different developmental stages.

PREMISE OF THE STUDY: Maca (Lepidium meyenii; Brassicaceae) has been cultivated by Andeans for thousands of years as a food source and has been used for medicinal purposes. However, little is known about the mechanism underlying material accumulation during plant growth. METHODS: RNA-Seq technology was used to compare the transcriptome of black maca root at three developmental stages. Gene Ontology term enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were applied for the identification of pathways in which differentially expressed genes were significantly enriched. RESULTS: Trinity was used to de novo assemble the reads, and 120,664 unigenes were assembled. Of these, 71.53% of the unigenes were annotated based on BLAST. A total of 18,321 differentially expressed genes were observed. Gene Ontology term enrichment analysis found that the most highly represented pathway among the differentially expressed genes was for genes involved in starch and sucrose metabolism. We also found that genes involved in secondary metabolite biosynthesis, such as glucosinolate biosynthesis, were significantly enriched. DISCUSSION: The genes that were differentially expressed between developmental time points likely reflect both developmental pathways and responses to changes in the environment. As such, the transcriptome data in this study serve as a reference for subsequent mining of genes that are involved in the synthesis of important bioactive components in maca.

Roles played by invertase and gene expression in the development of the horn-shaped gall on leaves of Rhus chinensis.

The present study deals with the growth and development of the horn-shaped gall, which is induced by Schlechtendalia chinensis Bell. on leaves of Rhus chinensis Mill. The relationship between gall formers and their host plants was investigated by means of the activities of various invertases, the expressions of the cell wall invertase gene (INV2), and vacuolar invertase gene (INV3) during gall development. Our results show that the increase in the sink strength of the galls required cell wall invertase and vacuolar invertase, and that vacuolar invertase had a particular impact during the early development. In addition, vacuolar invertase activity was always significantly higher in galls than in leaves. However, ionically bound cell wall invertase showed a slightly significant increased activity level when compared with the leaves after galls had entered the fast growing period. This result indicates that vacuolar invertase is related to the rapid expansion of the galls, but ionically bound cell wall invertase is involved in the rapid growth of tissues. The enhanced activity of cell wall invertase and the expression of INV2 may be a plant response to a gall-induced stress. Cytoplasmic invertase that acts as a maintenance enzyme, or takes part in the production of secondary metabolites, was elevated when intracellular acid invertase activity decreased.