Brg1 coordinates multiple processes during retinogenesis and is a tumor suppressor in retinoblastoma.

Retinal development requires precise temporal and spatial coordination of cell cycle exit, cell fate specification, cell migration and differentiation. When this process is disrupted, retinoblastoma, a developmental tumor of the retina, can form. Epigenetic modulators are central to precisely coordinating developmental events, and many epigenetic processes have been implicated in cancer. Studying epigenetic mechanisms in development is challenging because they often regulate multiple cellular processes; therefore, elucidating the primary molecular mechanisms involved can be difficult. Here we explore the role of Brg1 (Smarca4) in retinal development and retinoblastoma in mice using molecular and cellular approaches. Brg1 was found to regulate retinal size by controlling cell cycle length, cell cycle exit and cell survival during development. Brg1 was not required for cell fate specification but was required for photoreceptor differentiation and cell adhesion/polarity programs that contribute to proper retinal lamination during development. The combination of defective cell differentiation and lamination led to retinal degeneration in Brg1-deficient retinae. Despite the hypocellularity, premature cell cycle exit, increased cell death and extended cell cycle length, retinal progenitor cells persisted in Brg1-deficient retinae, making them more susceptible to retinoblastoma. ChIP-Seq analysis suggests that Brg1 might regulate gene expression through multiple mechanisms.

Enhanced Germicidal Efficacy by Co-Delivery of Validamycin and Hexaconazole with Methoxy Poly(ethylene glycol)-Poly(lactide-co-glycolide) Nanoparticles.

Co-delivery system has been proposed in pharmaceutical field aim to synergistic treatments. The combination formulation is also important in traditional pesticides formulations based on the low pest resistance risk and wide fungicidal spectrum. However, co-delivery nanoparticles (NPs) tend to be more environmentally friendly for the sustained-release behaviour and none of toxic organic solvents or dusts. Hence, we constructed co-delivery NPs which could delivery two kinds of pesticides, which function was similar with pesticides combination formulation. The co-delivery NPs of validamycin and hexaconazole were prepared with the amphiphilic copolymer methoxy poly(ethylene glycol)- poly(lactide-co-glycolide) (mPEG-PLGA) used an improved double emulsion method. The chemical structure of mPEG-PLGA copolymer was confirmed using fourier transform infrared spectroscopy (FT-IR), and nuclear magnetic resonance spectroscopy (NMR). The co-delivery NPs all exhibited good size distribution and held sustained-release property. Germicidal efficacy of the co-delivery NPs against Rhizoctonia cerealis was also studied. The germicidal efficacy of co-delivery NPs against Rhizoctonia cerealis was better than that of traditional pesticides formulation. In addition, co-delivery NPs showed a lasting impact against Rhizoctonia cerealis.

Size-Dependent Effect of Prochloraz-Loaded mPEG-PLGA Micro- and Nanoparticles.

As a controlled release formulation, polymer-based pesticide particle, provide an effective approach to achieve the target crop sites of increasing the pesticide utilization and reducing side effects. The particle size impacts on the dispersibility, pesticide loading content, control effect, etc. It is essential to investigate size-dependent effect. Hence, size-dependent effect of polymer-based pesticide particle was studied systematically in this paper. The biodegradable mPEG-PLGA copolymer with suitable molecular weight (45 KDa) was selected as carrier. Prochloraz-loaded mPEG-PLGA particles with different sizes (190.7 nm, 708.8 nm and 3980.0 nm) were constructed by emulsion/solvent evaporation method based on the same carrier. With the constant mass ratio of copolymer/prochloraz, as the particle size became large, the prochloraz loading content increased, and prochloraz released speed decreased. All prochloraz-loaded particles showed a sustained-release process and sustained impact against the Fusarium graminearum. Among the prochloraz-loaded mPEG-PLGA particles, the 190.7 nm particles exhibited the best germicidal efficacy in two weeks. Hence, the smaller size particles hold a better control efficacy in short time.

Pharmacokinetics and efficacy of the spleen tyrosine kinase inhibitor r406 after ocular delivery for retinoblastoma.

PURPOSE: Retinoblastoma is a childhood cancer of the retina. Clinical trials have shown that local delivery of broad spectrum chemotherapeutic agents is efficacious. Recent studies characterizing the genomic and epigenomic landscape of retinoblastoma identified spleen tyrosine kinase (SYK) as a promising candidate for targeted therapy. The purpose of this study was to conduct preclinical testing of the SYK antagonist R406 to evaluate it as a candidate for retinoblastoma treatment. METHODS: The efficacy of the SYK antagonist R406 delivered locally in a human orthotopic xenograft mouse model of retinoblastoma was tested. Intraocular exposure of R406 was determined for various routes and formulations. RESULTS: There was no evidence of efficacy for subconjunctival. R406. Maximal vitreal concentration was 10-fold lower than the minimal concentration required to kill retinoblastoma cells in vitro. Dosage of R406 subconjunctivally from emulsion or suspension formulations, direct intravitreal injection of the soluble prodrug of R406 (R788), and repeated topical administration of R406 all increased vitreal exposure, but failed to reach the exposure required for retinoblastoma cell death in culture. CONCLUSION: Taken together, these data suggest that R406 is not a viable clinical candidate for the treatment of retinoblastoma. This study highlights the importance of pharmacokinetic testing of molecular targeted retinoblastoma therapeutics.

Retinoblastoma (Rb) regulates laminar dendritic arbor reorganization in retinal horizontal neurons.

Neuronal differentiation with respect to the acquisition of synaptic competence needs to be regulated precisely during neurogenesis to ensure proper formation of circuits at the right place and time in development. This regulation is particularly important for synaptic triads among photoreceptors, horizontal cells (HCs), and bipolar cells in the retina, because HCs are among the first cell types produced during development, and bipolar cells are among the last. HCs undergo a dramatic transition from vertically oriented neurites that form columnar arbors to overlapping laminar dendritic arbors with differentiation. However, how this process is regulated and coordinated with differentiation of photoreceptors and bipolar cells remains unknown. Previous studies have suggested that the retinoblastoma (Rb) tumor suppressor gene may play a role in horizontal cell differentiation and synaptogenesis. By combining genetic mosaic analysis of individual synaptic triads with neuroanatomic analyses and multiphoton live imaging of developing HCs, we found that Rb plays a cell-autonomous role in the reorganization of horizontal cell neurites as they differentiate. Aberrant vertical processes in Rb-deficient HCs form ectopic synapses with rods in the outer nuclear layer but lack bipolar dendrites. Although previous reports indicate that photoreceptor abnormalities can trigger formation of ectopic synapses, our studies now demonstrate that defects in a postsynaptic partner contribute to the formation of ectopic photoreceptor synapses in the mammalian retina.

Antibiotic resistance of Helicobacter pylori isolated in the Southeast Coastal Region of China.

BACKGROUND: The resistance of Helicobacter pylori (H. pylori) to antibiotics is increasing worldwide, lowering its efficacy in current eradication therapies. This study evaluated H. pylori resistance to antibiotics in the southeast coastal region of China and suggests appropriate alternatives. MATERIALS AND METHODS: Seventeen thousand seven hundred and thirty one H. pylori strains were collected from eight areas of two provinces in coastal southeast China from 2010 to 2012. The resistance of these strains to six antibiotics was tested using the agar dilution method. RESULTS: The resistance rates to clarithromycin, metronidazole, levofloxacin, amoxicillin, gentamicin and furazolidone were 21.5, 95.4, 20.6, 0.1, 0.1 and 0.1%, respectively. Double, triple and quadruple antibacterial resistant percentages were 25.5, 7.5 and 0.1%, respectively. A positive association between the resistance to levofloxacin and to clarithromycin was found, but there was a negative correlation in the resistances to levofloxacin and to metronidazole. CONCLUSIONS: The prevalence of H. pylori resistance to clarithromycin, metronidazole, levofloxacin and multiple antibiotics in coastal southeast China is high. Choice of therapy should be individualized based on a susceptibility test in this region of the country.

Rb regulates proliferation and rod photoreceptor development in the mouse retina.

The retinoblastoma protein (Rb) regulates proliferation, cell fate specification and differentiation in the developing central nervous system (CNS), but the role of Rb in the developing mouse retina has not been studied, because Rb-deficient embryos die before the retinas are fully formed. We combined several genetic approaches to explore the role of Rb in the mouse retina. During postnatal development, Rb is expressed in proliferating retinal progenitor cells and differentiating rod photoreceptors. In the absence of Rb, progenitor cells continue to divide, and rods do not mature. To determine whether Rb functions in these processes in a cell-autonomous manner, we used a replication-incompetent retrovirus encoding Cre recombinase to inactivate the Rb1(lox) allele in individual retinal progenitor cells in vivo. Combined with data from studies of conditional inactivation of Rb1 using a combination of Cre transgenic mouse lines, these results show that Rb is required in a cell-autonomous manner for appropriate exit from the cell cycle of retinal progenitor cells and for rod development.

Cobalt-Phosphate (Co-Pi)-Modified WO3 Photoanodes for Performance-Enhanced Photoelectrochemical Wastewater Degradation.

Photocatalytic technology, with features of wide applicability, mild reaction conditions and sunlight availability, satisfies the requirements of "green chemistry". As the star photoanode material for photoelectrochemical catalysis, WO3 has a suitable band gap of 2.8 eV and a strong oxidation capacity, as well as displaying great potential in organic wastewater degradation. However, its performance is usually hindered by competition with water oxidation to generate peroxides, rapid charge complexation caused by surface defect sites, and so on. Herein, WO3 films modified with cobalt-phosphate (Co-Pi/WO3) film were prepared and involved in photocatalytic organic wastewater degradation. A degradation rate constant of 0.63311 h-1 was obtained for Co-Pi/WO3, which was much higher than that of WO3, 10.23 times that of direct photocatalysis (DP) and 23.99 times that of electrocatalysis (EC). After three cycles of degradation, the film can maintain a relatively good level of stability and a degradation efficiency of 93.79%.

Inhibition of CARM1-Mediated Methylation of ACSL4 Promotes Ferroptosis in Colorectal Cancer.

Ferroptosis, which is caused by iron-dependent accumulation of lipid peroxides, is an emerging form of regulated cell death and is considered a potential target for cancer therapy. However, the regulatory mechanisms underlying ferroptosis remain unclear. This study defines a distinctive role of ferroptosis. Inhibition of CARM1 can increase the sensitivity of tumor cells to ferroptosis inducers in vitro and in vivo. Mechanistically, it is found that ACSL4 is methylated by CARM1 at arginine 339 (R339). Furthermore, ACSL4 R339 methylation promotes RNF25 binding to ACSL4, which contributes to the ubiquitylation of ACSL4. The blockade of CARM1 facilitates ferroptosis and effectively enhances ferroptosis-associated cancer immunotherapy. Overall, this study demonstrates that CARM1 is a critical contributor to ferroptosis resistance and highlights CARM1 as a candidate therapeutic target for improving the effects of ferroptosis-based antitumor therapy.

Hypso- or bathochromic phosphorescent mechanochromic mononuclear Cu(I) complexes with a bis(2-diphenylphosphinophenyl)ether auxiliary ligand.

Six phosphorescence-emitting metal-organic mononuclear Cu(I) complexes, namely four quinoline-containing three-coordinate Cu(I) complexes and two N-heterocyclic carbene-containing four-coordinate Cu(I) complexes, have been successfully developed and fully characterized. All these Cu(I) complexes include the same bis(2-diphenylphosphinophenyl)ether bidentate auxiliary ligand. Significantly, four-coordinate Cu(I) complexes 1 and 2 display typical aggregation-induced emission phenomena. Their solid samples of luminogenic complexes 1-6 emit a variety of different phosphorescence. Furthermore, solid-state phosphorescence of these Cu(I) complexes can be effectively manipulated by external mechanical force. Remarkably, luminophores 1, 2 and 5 exhibit blue-shifted mechanoluminochromism responses, while luminophores 3, 4 and 6 present red-shifted mechanoluminochromism characteristics. All of the observed mechano-responsive phosphorescence changes of solids 1-6 are reversible by the method of solvent fuming. Powder X-ray diffraction results confirm that the reversible mechanically induced phosphorescence changes of complexes 1-6 are due to the mutual transformation of ordered crystalline and metastable amorphous states.

Inactivation of the p53 pathway in retinoblastoma.

Most human tumours have genetic mutations in their Rb and p53 pathways, but retinoblastoma is thought to be an exception. Studies suggest that retinoblastomas, which initiate with mutations in the gene retinoblastoma 1 (RB1), bypass the p53 pathway because they arise from intrinsically death-resistant cells during retinal development. In contrast to this prevailing theory, here we show that the tumour surveillance pathway mediated by Arf, MDM2, MDMX and p53 is activated after loss of RB1 during retinogenesis. RB1-deficient retinoblasts undergo p53-mediated apoptosis and exit the cell cycle. Subsequently, amplification of the MDMX gene and increased expression of MDMX protein are strongly selected for during tumour progression as a mechanism to suppress the p53 response in RB1-deficient retinal cells. Our data provide evidence that the p53 pathway is inactivated in retinoblastoma and that this cancer does not originate from intrinsically death-resistant cells as previously thought. In addition, they support the idea that MDMX is a specific chemotherapeutic target for treating retinoblastoma.

Construction of three-dimensional cobalt sulfide/multi-heteroatom co-doped porous carbon as an efficient trifunctional electrocatalyst.

Exploring cost-effective non-precious metal electrocatalysts is vital for the large-scale application of clean energy conversion devices (i.e., fuel cells, metal-air batteries and water electrolysers). Herein, we present the construction of a three-dimensional cobalt sulfide/multi-heteroatom co-doped carbon composite as a trifunctional electrocatalyst for the oxygen reduction reaction (ORR), oxygen evolution reaction (OER) and hydrogen evolution reaction (HER) through one-step sulfidation of zeolitic-imidazolate frameworks (ZIFs) using sulfur powder as a sulfur source. By virtue of the distinct periodic metal-nitrogen coordination structure and the abundant micropores within the ZIF precursor, sub-10 nm Co9S8 nanoparticles (NPs) are homogenously anchored on a Co, S and N multi-heteroatom co-doped carbon framework with a large specific surface area that exposes sufficient reactive sites for these electrocatalytic reactions. The optimized Co9S8/CoNSC exhibits outstanding ORR, OER and HER performance, comparable or even superior to those of commercial Pt/C and RuO2. The small Co9S8 NPs and Co-Nx species embedded in the carbon matrix cooperatively catalyze the OER and ORR, while the HER catalysis is mainly contributed by Co9S8 NPs. Furthermore, the Co9S8/CoNSC shows outstanding anti-poisoning capability towards sulfur species during ORR catalysis with no obvious activity degradation observed in 0.1 M KOH containing 50 µM SO32- species, significantly outperforming commercial Pt/C. The assembled rechargeable Zn-air battery using the Co9S8/CoNSC as a cathode shows a high power density (150 mW cm-2) and the assembled water electrolyzer only requires 1.585 V at a current density of 10 mA cm-2 when using this material as an anode and a cathode. This work provides an effective strategy to design and synthesize efficient, durable and anti-poisoning cobalt chalcogenide-based trifunctional electrocatalysts for the large-scale application of clean energy conversion devices.

Surface Engineering of Carbon-Supported Platinum as a Route to Electrocatalysts with Superior Durability and Activity for PEMFC Cathodes.

Hydrogen fuel cells are regarded as a promising new carbon mitigation strategy to realize carbon neutrality. The exploitation of robust and efficient cathode catalysts is thus vital to the commercialization of proton exchange membrane fuel cells (PEMFCs). Herein, we demonstrate a facile and scalable surface engineering route to achieve superior durability and high activity of a Pt-based material as a PEMFC cathode catalyst through a controllable liquid-phase reduction approach. The proposed surface engineering strategy by modifying Pt/C reduces the oxygen content on the carbon support and also decreases the surface defects on Pt nanoparticles (NPs), which effectively alleviate the corrosion of carbon and inhibit the detachment, agglomeration, and growth of Pt NPs. The resulting catalyst exhibits superior durability after a 10,000 potential cycling test in an acid electrolyte─outperforming commercial Pt/C. Moreover, the catalyst also demonstrates an improved oxygen reduction reaction (ORR) activity in comparison to commercial Pt/C by virtue of the high content of metallic Pt and the weakened Pt-OH bonding that releases more Pt active sites for ORR catalysis. Most importantly, the developed catalyst shows outstanding PEMFC performance and excellent long-term durability over 50 h of a constant-current test and 100 h of a load-cycling operation. This effective route provides a new avenue for exploiting robust Pt-based catalysts with superior activity in practical applications of PEMFCs.

Intriguing H2S Tolerance of the PtRu Alloy for Hydrogen Oxidation Catalysis in PEMFCs: Weakened Pt-S Binding with Slower Adsorption Kinetics.

High quality of hydrogen is the key to the long lifetime of proton-exchange membrane fuel cell (PEMFC) vehicles, while trace H2S impurities in hydrogen significantly affect their durability and fuel expense. Herein, we demonstrate a robust PtRu alloy catalyst with an intriguing H2S tolerance as the PEMFC anode, showing a stronger antipoisoning capability toward hydrogen oxidation reaction compared with the Pt/C anode. The PtRu/C-based single PEMFC shows approximately 14.3% loss of cell voltage after 3 h operation with 1 ppm of H2S in hydrogen, significantly lower than that of Pt/C-based PEMFCs (65%). By adopting PtRu/C as the anode, the H2S limit in hydrogen can be increased to 1.7 times that of the Pt/C anode, assuming that the PEMFC runs for 5000 h, which is conductive for the cost reduction of hydrogen purification. The three-electrode electrochemical test indicates that PtRu/C exhibits a slower adsorption kinetics toward S2- species with poisoning rates of 0.02782, 0.02982, and 0.03682 min-1 at temperatures of 25, 35, and 45 °C, respectively, all lower than those of Pt/C. X-ray absorption fine structure spectra indicate the weakened Pt-S binding for PtRu/C in comparison to Pt/C with a longer Pt-S bond length. Density functional theory calculation analyses reveal that adsorption energy of sulfur on the Pt surface was reduced for PtRu/C, showing 1-10% decrease at different Pt sites for (111), (110), and (100) planes, which is ascribed to the downshifted Pt d-band center caused by the ligand and strain effects due to the introduction of second metallic Ru. This work provides a valuable guide for the development of the H2S-tolerant catalysts for long-term application of PEMFCs.

Identification of a modular super-enhancer in murine retinal development.

Super-enhancers are expansive regions of genomic DNA comprised of multiple putative enhancers that contribute to the dynamic gene expression patterns during development. This is particularly important in neurogenesis because many essential transcription factors have complex developmental stage- and cell-type specific expression patterns across the central nervous system. In the developing retina, Vsx2 is expressed in retinal progenitor cells and is maintained in differentiated bipolar neurons and Müller glia. A single super-enhancer controls this complex and dynamic pattern of expression. Here we show that deletion of one region disrupts retinal progenitor cell proliferation but does not affect cell fate specification. The deletion of another region has no effect on retinal progenitor cell proliferation but instead leads to a complete loss of bipolar neurons. This prototypical super-enhancer may serve as a model for dissecting the complex gene expression patterns for neurogenic transcription factors during development. Moreover, it provides a unique opportunity to alter expression of individual transcription factors in particular cell types at specific stages of development. This provides a deeper understanding of function that cannot be achieved with traditional knockout mouse approaches.

[Clinical effect of cortical incision with rapid maxillary expansion to assist orthodontic-orthognathic treatment of skeletal Class â ¢ malocclusion].

PURPOSE: To investigate the clinical effect of rapid maxillary expansion with cortical osteotomy combined with orthodontic- orthognathic treatment for skeletal Class Ⅲ malocclusion. METHODS: A total of 84 patients with skeletal Class Ⅲ malocclusion admitted to Jining Dental Hospital from March 2018 to May 2020 were randomly divided into experimental group and control group, with 42 cases in each group. The control group was treated with orthodontic-orthognathic treatment, while the experimental group was treated with orthodontic-orthognathic treatment with rapid maxillary arch expansion by cortical incision. The time of closing gap, alignment time, maxillary first molar and maxillary central incisor tooth movement distance in the sagittal direction were compared between the two groups. Before treatment and 4 weeks after treatment, the vertical distance from the end of the upper central incisor edge to the horizontal plane(U1I-HP), the distance from the apex of the upper central incisor to the coronal plane(U1I-CP), the vertical distance from the edge of the upper pressure groove to the coronal plane(Sd-CP), the vertical distance from the upper alveolar seat point to the horizontal plane(A-HP), the vertical distance from the point of the upper lip to the coronal plane(Ls-CP), and the vertical distance from the inferior point of nose to coronal plane(Sn-CP) were measured, and the relevant changes were calculated. During the treatment period, the complications of the two groups were compared. SPSS 20.0 software package was used for statistical analysis of the data. RESULTS: There was no significant difference in alignment time, A-HP change, Sn-CP change, maxillary first molar movement distance and maxillary central incisor movement distance between the two groups (P＞0.05). The closing interval in the experimental group was significantly shorter than that of the control group (P＜0.05). The changes of U1I-HP, U1I-CP, Sd-CP, and Ls-CP in the experimental group were significantly higher than those in the control group (P＜0.05). There was no significant difference in the incidence of complications between the two groups during treatment (P＞0.05). CONCLUSIONS: Rapid maxillary expansion of cortical incision assisted orthodontic-orthognathic treatment of skeletal Class Ⅲ malocclusion patients can shorten the closing gap time and improve the treatment effect, which has no obvious influence on the sagittal direction of the teeth.

Whole-body physiologically based pharmacokinetic model for nutlin-3a in mice after intravenous and oral administration.

Nutlin-3a is an MDM2 inhibitor that is under investigation in preclinical models for a variety of pediatric malignancies, including retinoblastoma, rhabdomyosarcoma, neuroblastoma, and leukemia. We used physiologically based pharmacokinetic (PBPK) modeling to characterize the disposition of nutlin-3a in the mouse. Plasma protein binding and blood partitioning were assessed by in vitro studies. After intravenous (10 and 20 mg/kg) and oral (50, 100, and 200 mg/kg) dosing, tissue concentrations of nutlin-3a were determined in plasma, liver, spleen, intestine, muscle, lung, adipose, bone marrow, adrenal gland, brain, retina, and vitreous fluid. The PBPK model was simultaneously fit to all pharmacokinetic data using NONMEM. Nutlin-3a exhibited nonlinear binding to murine plasma proteins, with the unbound fraction ranging from 0.7 to 11.8%. Nutlin-3a disposition was characterized by rapid absorption with peak plasma concentrations at approximately 2 h and biphasic elimination consistent with a saturable clearance process. The final PBPK model successfully described the plasma and tissue disposition of nutlin-3a. Simulations suggested high bioavailability, rapid attainment of steady state, and little accumulation when administered once or twice daily at dosages up to 400 mg/kg. The final model was used to perform simulations of unbound tissue concentrations to determine which dosing regimens are appropriate for preclinical models of several pediatric malignancies.

Retinoblastoma from human stem cell-derived retinal organoids.

Retinoblastoma is a childhood cancer of the developing retina that initiates with biallelic inactivation of the RB1 gene. Children with germline mutations in RB1 have a high likelihood of developing retinoblastoma and other malignancies later in life. Genetically engineered mouse models of retinoblastoma share some similarities with human retinoblastoma but there are differences in their cellular differentiation. To develop a laboratory model of human retinoblastoma formation, we make induced pluripotent stem cells (iPSCs) from 15 participants with germline RB1 mutations. Each of the stem cell lines is validated, characterized and then differentiated into retina using a 3-dimensional organoid culture system. After 45 days in culture, the retinal organoids are dissociated and injected into the vitreous of eyes of immunocompromised mice to support retinoblastoma tumor growth. Retinoblastomas formed from retinal organoids made from patient-derived iPSCs have molecular, cellular and genomic features indistinguishable from human retinoblastomas. This model of human cancer based on patient-derived iPSCs with germline cancer predisposing mutations provides valuable insights into the cellular origins of this debilitating childhood disease as well as the mechanism of tumorigenesis following RB1 gene inactivation.

Neuronal differentiation and synaptogenesis in retinoblastoma.

Retinoblastomas initiate in the developing retina in utero and are diagnosed during the first few years of life. We have recently generated a series of knockout mouse models of retinoblastoma that recapitulate the timing, location, and progression of human retinoblastoma. One of the most important benefits of these preclinical models is that we can study the earliest stages of tumor initiation and expansion. This is not possible in human retinoblastoma because tumors initiate in utero and are not diagnosed until they are at an advanced stage. We found that mouse retinoblastoma cells exhibit a surprising degree of differentiation, which has not been previously reported for any neural tumor. Early-stage mouse retinoblastoma cells express proteins found normally in retinal plexiform layers. They also extend neurites and form synapses. All of these features, which were characterized by immunostaining, Golgi-Cox staining, scanning electron microscopy, and transmission electron microscopy, suggest that mouse retinoblastoma cells resemble amacrine/horizontal cells from the retina. As late-stage retinoblastoma cells expand and invade the surrounding tissue, they lose their differentiated morphology and become indistinguishable from human retinoblastomas. Taken together, our data suggest that neuronal differentiation is a hallmark of early-stage retinoblastoma and is lost as cells become more aggressive and invasive. We also show that rosette formation is not a hallmark of retinoblastoma differentiation, as previously believed. Instead, rosette formation reflects extensive cell-cell contacts between retinoblastoma cells in both early-stage (differentiated) and late-stage (dedifferentiated) tumors.

Nucleome Dynamics during Retinal Development.

More than 8,000 genes are turned on or off as progenitor cells produce the 7 classes of retinal cell types during development. Thousands of enhancers are also active in the developing retinae, many having features of cell- and developmental stage-specific activity. We studied dynamic changes in the 3D chromatin landscape important for precisely orchestrated changes in gene expression during retinal development by ultra-deep in situ Hi-C analysis on murine retinae. We identified developmental-stage-specific changes in chromatin compartments and enhancer-promoter interactions. We developed a machine learning-based algorithm to map euchromatin and heterochromatin domains genome-wide and overlaid it with chromatin compartments identified by Hi-C. Single-cell ATAC-seq and RNA-seq were integrated with our Hi-C and previous ChIP-seq data to identify cell- and developmental-stage-specific super-enhancers (SEs). We identified a bipolar neuron-specific core regulatory circuit SE upstream of Vsx2, whose deletion in mice led to the loss of bipolar neurons.