Key transcription factors influence the epigenetic landscape to regulate retinal cell differentiation.

How the diverse neural cell types emerge from multipotent neural progenitor cells during central nervous system development remains poorly understood. Recent scRNA-seq studies have delineated the developmental trajectories of individual neural cell types in many neural systems including the neural retina. Further understanding of the formation of neural cell diversity requires knowledge about how the epigenetic landscape shifts along individual cell lineages and how key transcription factors regulate these changes. In this study, we dissect the changes in the epigenetic landscape during early retinal cell differentiation by scATAC-seq and identify globally the enhancers, enriched motifs, and potential interacting transcription factors underlying the cell state/type specific gene expression in individual lineages. Using CUT&Tag, we further identify the enhancers bound directly by four key transcription factors, Otx2, Atoh7, Pou4f2 and Isl1, including those dependent on Atoh7, and uncover the sequential and combinatorial interactions of these factors with the epigenetic landscape to control gene expression along individual retinal cell lineages such as retinal ganglion cells (RGCs). Our results reveal a general paradigm in which transcription factors collaborate and compete to regulate the emergence of distinct retinal cell types such as RGCs from multipotent retinal progenitor cells (RPCs).

Two new genetically modified mouse alleles labeling distinct phases of retinal ganglion cell development by fluorescent proteins.

BACKGROUND: During development, all retinal cell types arise from retinal progenitor cells (RPCs) in a step-wise fashion. Atoh7 and Pou4f2 mark, and function in, two phases of retinal ganglion cell (RGC) genesis; Atoh7 functions in a subpopulation of RPCs to render them competent for the RGC fate, whereas Pou4f2 participates in RGC fate specification and RGC differentiation. Despite extensive research on their roles, the properties of the two phases represented by these two factors have not been well studied, likely due to the retinal cellular heterogeneity. RESULTS: In this report, we describe two novel knock-in mouse alleles, Atoh7zsGreenCreERT2 and Pou4f2FlagtdTomato , which labeled retinal cells in the two phases of RGC development by fluorescent proteins. Also, the Atoh7zsGreenCreERT2 allele allowed for indirect labeling of RGCs and other cell types upon tamoxifen induction in a dose-dependent manner. Further, these alleles could be used to purify retinal cells in the different phases by fluorescence assisted cell sorting (FACS). Single cell RNA-seq analysis of purified cells from Atoh7zsGreenCreERT2 retinas further validated that this allele labeled both transitional/competent RPCs and their progenies including RGCs. CONCLUSIONS: Thus, these two alleles are very useful tools for studying the molecular and genetic mechanisms underlying RGC formation.

[Influence of the Protamine Sulfate on Endocytosis and Intracellular Lysosome Escape of DNA Nanostructure].

OBJECTIVE: To investigate the influence of the protamine sulfate on endocytosis and intracellular stability of tetrahedral framework nucleic acid (tFNA). METHODS: Articular cartilage cells were collected from 3-day-old C57BL mice. Cells at passage 1-2 were used in the experiments. 4 single-strand DNAs (S1 was marked by Cy5) were utilized to synthesize tFNAs via annealing process and ultrafiltration for purification. High-performance capillary electrophoresis (HPCE) was used to verify synthesis of tFNAs and transmission electron microscope was used to photo morphological characteristics. The 1 mg/mL protamine sulfate solution was slowly dropped into newly synthesized tFNAs (N/P=5/1). Then, Zeta potential was detected. Cells were treated with 100 nmol/L tFNAs with protamine sulfate in Dulbecco's Modified Eagle's medium (DMEM) (Exp.1), 100 nmol/L tFNAs in DMEM (Exp.2), and DMEM (Control), respectively. Flow cytometry was used to quantitatively detect intracellular Cy5 fluorescence after 6 h and 12 h treatments. Immunofluorescence staining was used to qualitatively observe internalized Cy5 fluorescence after 12 h treatment by laser confocal microscope. Lysosome of living cells were stained with lysosome probe. Colocalization between lysosome and tFNAs was observed by laser confocal microscope. RESULTS: After incubating protamine sulfate, negative potential was transformed into positive one ( (-1.567±0.163) mV to (4.700±0.484) mV). The fluorescence intensity of tFNAs in the Exp.1 group was higher than that of the Exp.2 group in 6 h and 12 h ( P<0.05). This was consistent with the results of immunofluorescence staining after 12 h. Colocalization of Cy5 fluorescence and lysosome in the Exp.1 group was more rare than that in the Exp.2 group at 6 h and 12 h. Furthermore, a large amount of Cy5 fluorescence was still seen in the Exp.1 group at 12 h, while Cy5 fluorescence of the Exp.2 group was less. CONCLUSION: Protamine sulfate can effectively enhance endocytosis, and to some extent it can achieve lysosome escape of tFNAs.

GMDTC Chelating Agent Attenuates Cisplatin-Induced Systemic Toxicity without Affecting Antitumor Efficacy.

Cisplatin is a platinum-based chemotherapeutic drug widely used in the treatment of various cancers such as testicular, ovarian, lung, bladder, and cervical cancers. However, its use and the dosage range applied have been limited by severe side effects (e.g., nephrotoxicity and ototoxicity) and by the development of resistance to cisplatin in patients during treatment. Metal chelators have shown promising potential in overcoming these problems often associated with platinum drugs. Previously, a new chelating agent, sodium (S)-2-(dithiocarboxylato((2S,3R,4R,5R)-2,3,4,5,6-pentahydroxyhexyl)amino)-4(methylthio)butanoate (GMDTC), was developed. In this study, we examined the effect of GMDTC in modifying cisplatin-induced toxicities following in vitro and in vivo exposures. GMDTC treatment dramatically reduced cisplatin-induced apoptosis and cytotoxicity in HK2 cells by decreasing the amount of intracellular platinum. In the 4T1 breast cancer mouse model, GMDTC reduced cisplatin-induced nephrotoxicity by reducing cisplatin deposition in the kidney. GMDTC attenuated cisplatin-induced elevations in blood urea nitrogen and plasma creatinine, ameliorated renal tubular dilation and vacuolation, and prevented necrosis of glomeruli and renal tubular cells. GMDTC also inhibited cisplatin-induced ototoxicity as shown by improved hearing loss which was assessed using the auditory brainstem response test. Furthermore, GMDTC attenuated cisplatin-induced hematotoxicity and hepatotoxicity. Importantly, co-treatment of cisplatin with GMDTC did not affect cisplatin antitumor efficacy. Tumor growth, size, and metastasis were all comparable between the cisplatin only and cisplatin-GMDTC co-treatment groups. In conclusion, the current study suggests that GMDTC reduces cisplatin-induced systemic toxicity by preventing the accumulation and assisting in the removal of intracellular cisplatin, without compromising cisplatin therapeutic activity. These results support the development of GMDTC as a chemotherapy protector and rescue agent to overcome the toxicity of and resistance to platinum-based antineoplastic drugs.

Mapping dynamic histone modification patterns during arsenic-induced malignant transformation of human bladder cells.

Arsenic is a known potent risk factor for bladder cancer. Increasing evidence suggests that epigenetic alterations, e.g., DNA methylation and histones posttranslational modifications (PTMs), contribute to arsenic carcinogenesis. Our previous studies have demonstrated that exposure of human urothelial cells (UROtsa cells) to monomethylarsonous acid (MMAIII), one of arsenic active metabolites, changes the histone acetylation marks across the genome that are correlated with MMAIII-induced UROtsa cell malignant transformation. In the current study, we employed a high-resolution and high-throughput liquid chromatography tandem mass spectrometry (LC-MS/MS) to identify and quantitatively measure various PTM patterns during the MMAIII-induced malignant transformation. Our data showed that MMAIII exposure caused a time-dependent increase in histone H3 acetylation on lysine K4, K9, K14, K18, K23, and K27, but a decrease in acetylation on lysine K5, K8, K12, and K16 of histone H4. Consistent with this observation, H3K18ac was increased while H4K8ac was decreased in the leukocytes collected from people exposed to high concentrations of arsenic compared to those exposed to low concentrations. MMAIII was also able to alter histone methylation patterns: MMAIII transformed cells experienced a loss of H3K4me1, and an increase in H3K9me1 and H3K27me1. Collectively, our data shows that arsenic exposure causes dynamic changes in histone acetylation and methylation patterns during arsenic-induced cancer development. Exploring the genomic location of the altered histone marks and the resulting aberrant expression of genes will be of importance in deciphering the mechanism of arsenic-induced carcinogenesis.

Taxonomic and Functional Analyses of the Supragingival Microbiome from Caries-Affected and Caries-Free Hosts.

Caries is one of the most prevalent and costly infectious diseases affecting humans of all ages. It is initiated by cariogenic supragingival dental plaques forming on saliva-coated tooth surfaces, yet the etiology remains elusive. To determine which microbial populations may predispose a patient to caries, we report here an in-depth and comprehensive view of the microbial community associated with supragingival dental plaque collected from the healthy teeth of caries patients and healthy adults. We found that microbial communities from caries patients had a higher evenness and inter-individual variations but simpler ecological networks compared to healthy controls despite the overall taxonomic structure being similar. Genera including Selenomonas, Treponema, Atopobium, and Bergeriella were distributed differently between the caries and healthy groups with disturbed co-occurrence patterns. In addition, caries and healthy subjects carried different Treponema, Atopobium, and Prevotella species. Moreover, distinct populations of 13 function genes involved in organic acid synthesis, glycan biosynthesis, complex carbohydrate degradation, amino acid synthesis and metabolism, purine and pyrimidine metabolism, isoprenoid biosynthesis, lipid metabolism, and co-factor biosynthesis were present in each of the healthy and caries groups. Our results suggested that the fundamental differences in dental plaque ecology partially explained the patients' susceptibility to caries, and could be used for caries risk prediction in the future.

Comparison of hematological alterations and markers of B-cell activation in workers exposed to benzene, formaldehyde and trichloroethylene.

Benzene, formaldehyde (FA) and trichloroethylene (TCE) are ubiquitous chemicals in workplaces and the general environment. Benzene is an established myeloid leukemogen and probable lymphomagen. FA is classified as a myeloid leukemogen but has not been associated with non-Hodgkin lymphoma (NHL), whereas TCE has been associated with NHL but not myeloid leukemia. Epidemiologic associations between FA and myeloid leukemia, and between benzene, TCE and NHL are, however, still debated. Previously, we showed that these chemicals are associated with hematotoxicity in cross-sectional studies of factory workers in China, which included extensive personal monitoring and biological sample collection. Here, we compare and contrast patterns of hematotoxicity, monosomy 7 in myeloid progenitor cells (MPCs), and B-cell activation biomarkers across these studies to further evaluate possible mechanisms of action and consistency of effects with observed hematologic cancer risks. Workers exposed to benzene or FA, but not TCE, showed declines in cell types derived from MPCs, including granulocytes and platelets. Alterations in lymphoid cell types, including B cells and CD4+ T cells, and B-cell activation markers were apparent in workers exposed to benzene or TCE. Given that alterations in myeloid and lymphoid cell types are associated with hematological malignancies, our data provide biologic insight into the epidemiological evidence linking benzene and FA exposure with myeloid leukemia risk, and TCE and benzene exposure with NHL risk.

Chromosome-wide aneuploidy study of cultured circulating myeloid progenitor cells from workers occupationally exposed to formaldehyde.

Formaldehyde (FA) is an economically important industrial chemical to which millions of people worldwide are exposed environmentally and occupationally. Recently, the International Agency for Cancer Research concluded that there is sufficient evidence that FA causes leukemia, particularly myeloid leukemia. To evaluate the biological plausibility of this association, we employed a chromosome-wide aneuploidy study approach, which allows the evaluation of aneuploidy and structural chromosome aberrations (SCAs) of all 24 chromosomes simultaneously, to analyze cultured myeloid progenitor cells from 29 workers exposed to relatively high levels of FA and 23 unexposed controls. We found statistically significant increases in the frequencies of monosomy, trisomy, tetrasomy and SCAs of multiple chromosomes in exposed workers compared with controls, with particularly notable effects for monosomy 1 [P = 6.02E-06, incidence rate ratio (IRR) = 2.31], monosomy 5 (P = 9.01E-06; IRR = 2.24), monosomy 7 (P = 1.57E-05; IRR = 2.17), trisomy 5 (P = 1.98E-05; IRR = 3.40) and SCAs of chromosome 5 (P = 0.024; IRR = 4.15). The detection of increased levels of monosomy 7 and SCAs of chromosome 5 is particularly relevant as they are frequently observed in acute myeloid leukemia. Our findings provide further evidence that leukemia-related cytogenetic changes can occur in the circulating myeloid progenitor cells of healthy workers exposed to FA, which may be a potential mechanism underlying FA-induced leukemogenesis.

Arsenic responsive microRNAs in vivo and their potential involvement in arsenic-induced oxidative stress.

Arsenic exposure is postulated to modify microRNA (miRNA) expression, leading to changes of gene expression and toxicities, but studies relating the responses of miRNAs to arsenic exposure are lacking, especially with respect to in vivo studies. We utilized high-throughput sequencing technology and generated miRNA expression profiles of liver tissues from Sprague Dawley (SD) rats exposed to various concentrations of sodium arsenite (0, 0.1, 1, 10 and 100mg/L) for 60days. Unsupervised hierarchical clustering analysis of the miRNA expression profiles clustered the SD rats into different groups based on the arsenic exposure status, indicating a highly significant association between arsenic exposure and cluster membership (p-value of 0.0012). Multiple miRNA expressions were altered by arsenic in an exposure concentration-dependent manner. Among the identified arsenic-responsive miRNAs, several are predicted to target Nfe2l2-regulated antioxidant genes, including glutamate-cysteine ligase (GCL) catalytic subunit (GCLC) and modifier subunit (GCLM) which are involved in glutathione (GSH) synthesis. Exposure to low concentrations of arsenic increased mRNA expression for Gclc and Gclm, while high concentrations significantly reduced their expression, which were correlated to changes in hepatic GCL activity and GSH level. Moreover, our data suggested that other mechanisms, e.g., miRNAs, rather than Nfe2l2-signaling pathway, could be involved in the regulation of mRNA expression of Gclc and Gclm post-arsenic exposure in vivo. Together, our findings show that arsenic exposure disrupts the genome-wide expression of miRNAs in vivo, which could lead to the biological consequence, such as an altered balance of antioxidant defense and oxidative stress.

Digital workflow in the design of individualized emergence profiles of implant restorations based on the contralateral tooth.

PURPOSE: To describe a novel digital design technique for creating an individualized emergence profile for implant restoration based on the contralateral tooth. METHODS: Cone beam computed tomography (CBCT) data were used to accurately obtain a three-dimensional (3D) model of the contralateral tooth, which was mirror-flipped to design the emergence profile. The emergence profile was further divided into critical and subcritical areas; the critical area precisely replicated the mirror-flipped 3D model, whereas the subcritical area featured a slight concavity on the buccal side, flatness on the lingual side, and slight convexity on the mesial and distal surfaces. Subsequently, a milling machine was used to fabricate healing abutments with individualized emergence profiles. The design of the definitive restoration completely duplicated the emergence profile of the individualized healing abutment and was fabricated using a milling machine. CONCLUSIONS: This technical procedure presents an alternative novel method for designing the emergence profiles of implant restorations, with the potential to improve esthetics and functions as well as to maintain the long-term stability of peri-implant soft and hard tissues.

Framework Nucleic Acids-Based VEGF Signaling Activating System for Angiogenesis: A Dual Stimulation Strategy.

Angiogenesis is crucial for tissue engineering, wound healing, and regenerative medicine. Nanomaterials constructed based on specific goals can be employed to activate endogenous growth factor-related signaling. In this study, based on the conventional single-stranded DNA self-assembly into tetrahedral framework nucleic acids (tFNAs), the Apt02 nucleic acid aptamer and dimethyloxallyl glycine (DMOG) small molecule are integrated into a complex via a template-based click chemistry reaction and toehold-mediated strand displacement reaction. Thus, being able to simulate the VEGF (vascular endothelial growth factor) function and stabilize HIF (hypoxia-inducible factor), a functional whole is constructed and applied to angiogenesis. Cellular studies demonstrate that the tFNAs-Apt02 complex (TAC) has a conspicuous affinity to human umbilical vein endothelial cells (HUVECs). Further incubation with DMOG yields the tFNAs-Apt02-DMOG complex (TACD), which promotes VEGF secretion, in vitro blood vessel formation, sprouting, and migration of HUVECs. Additionally, TACD enhances angiogenesis by upregulating the VEGF/VEGFR and HIF signaling pathways. Moreover, in a diabetic mouse skin defect repair process, TACD increases blood vessel formation and collagen deposition, therefore accelerating wound healing. The novel strategy simulating VEGF and stabilizing HIF promotes blood-vessel formation in vivo and in vitro and has the potential for broad applications in the vascularization field.

Alterations in serum immunoglobulin levels in workers occupationally exposed to trichloroethylene.

Trichloroethylene (TCE) has been associated with a variety of immunotoxic effects and may be associated with an increased risk of non-Hodgkin lymphoma (NHL). Altered serum immunoglobulin (Ig) levels have been reported in NHL patients and in animals exposed to TCE. Recently, we reported that occupational exposure to TCE is associated with immunosuppressive effects and immune dysfunction, including suppression of B-cell counts and activation, even at relatively low levels. We hypothesized that TCE exposure would also affect Ig levels in humans. We measured serum levels of IgG, IgM and IgE, by enzyme-linked immunosorbent assay, in TCE-exposed workers (n = 80) and unexposed controls (n = 45), matched by age and gender, in a cross-sectional, molecular epidemiology study of occupational exposure to TCE in Guangdong, China. Exposed workers had about a 17.5% decline in serum levels of IgG compared with unexposed controls (P = 0.0002). Similarly, serum levels of IgM were reduced by about 38% in workers exposed to TCE compared with unexposed controls (P < 0.0001). Serum levels of both IgG and IgM were significantly decreased in workers exposed to TCE levels below 12 p.p.m., the median exposure level. Adjustment for B-cell counts had minimal impact on our findings. IgE levels were not significantly different between exposed and control subjects. These results provide further evidence that TCE is immunotoxic at relatively low exposure levels and provide additional biologic plausibility for the reported association of TCE with NHL.

Chronic low level trimethyltin exposure and the risk of developing nephrolithiasis.

OBJECTIVES: Nephrolithiasis (kidney stones) is a common disease with the prevalence that is increasing globally. We previously found that trimethyltin (TMT), a by-product of plastic stabilisers, can inhibit the H(+)/K(+) ATPase activity in renal intercalated cells and alter urinary pH, which is a known risk factor for nephrolithiasis. In this study, we conducted a cross-sectional analysis to evaluate the impact of chronic low level occupational TMT exposure on nephrolithiasis. METHODS: This study included 216 healthy workers with TMT exposure and 119 workers as controls with no TMT exposure. All study participants were administered a questionnaire and underwent a routine clinical examination including an ultrasonographic screening for kidney stones. Exposures were assessed by measuring TMT concentrations in personal air samples, blood and urine. Logistic regression analysis was used to estimate the ORs and 95% CIs for the risk of kidney stones. RESULTS: TMT exposed workers had a higher prevalence of kidney stones (18.06%) in comparison with control workers (5.88%). High TMT concentrations in personal air samples, blood and urines were positively associated with increased prevalence of kidney stones in workers exposed to TMT compared with controls workers (p-trend values=0.005, 0.008 and 0.002, respectively). The length of employment in plants with elevated TMT levels (duration of the exposure) was significantly associated with the increased prevalence of kidney stones (p trend=0.001). The ORs were 2.66 for <3 years, 3.73 for 3-<10 years and 7.89 for 10+ years of employment compared with control workers. CONCLUSIONS: To our knowledge, this is the first report to demonstrate that occupational exposure to TMT is a potential risk factor for nephrolithiasis.

Occupational exposure to formaldehyde and alterations in lymphocyte subsets.

BACKGROUND: Formaldehyde is used in many occupational settings, most notably in manufacturing, health care, and embalming. Formaldehyde has been classified as a human carcinogen, but its mechanism of action remains uncertain. METHODS: We carried out a cross-sectional study of 43 formaldehyde-exposed workers and 51 unexposed age and sex-matched controls in Guangdong, China to study formaldehyde's early biologic effects. To follow up our previous report that the total lymphocyte count was decreased in formaldehyde-exposed workers compared with controls, we evaluated each major lymphocyte subset (i.e., CD4(+) T cells, CD8(+) T cells, natural killer [NK] cells, and B cells) and T cell lymphocyte subset (CD4(+) naïve and memory T cells, CD8(+) naïve and memory T cells, and regulatory T cells). Linear regression of each subset was used to test for differences between exposed workers and controls, adjusting for potential confounders. RESULTS: Total NK cell and T cell counts were about 24% (P = 0.037) and 16% (P = 0.0042) lower, respectively, among exposed workers. Among certain T cell subsets, decreased counts among exposed workers were observed for CD8(+) T cells (P = 0.026), CD8(+) effector memory T cells (P = 0.018), and regulatory T cells (CD4(+) FoxP3(+) : P = 0.04; CD25(+) FoxP3(+) : P = 0.008). CONCLUSIONS: Formaldehyde-exposed workers experienced decreased counts of NK cells, regulatory T cells, and CD8(+) effector memory T cells; however, due to the small sample size; these findings need to be confirmed in larger studies.

Elevated urinary levels of kidney injury molecule-1 among Chinese factory workers exposed to trichloroethylene.

Epidemiological studies suggest that trichloroethylene (TCE) exposure may be associated with renal cancer. The biological mechanisms involved are not exactly known although nephrotoxicity is believed to play a role. Studies on TCE nephrotoxicity among humans, however, have been largely inconsistent. We studied kidney toxicity in Chinese factory workers exposed to TCE using novel sensitive nephrotoxicity markers. Eighty healthy workers exposed to TCE and 45 comparable unexposed controls were included in the present analyses. Personal TCE exposure measurements were taken over a 2-week period before urine collection. Ninety-six percent of workers were exposed to TCE below the current US Occupational Safety and Health Administration permissible exposure limit (100 ppm 8h TWA), with a mean (SD) of 22.2 (35.9) ppm. Kidney injury molecule-1 (KIM-1) and Pi-glutathione S transferase (GST) alpha were elevated among the exposed subjects as compared with the unexposed controls with a strong exposure-response association between individual estimates of TCE exposure and KIM-1 (P < 0.0001). This is the first report to use a set of sensitive nephrotoxicity markers to study the possible effects of TCE on the kidneys. The findings suggest that at relatively low occupational exposure levels a toxic effect on the kidneys can be observed. This finding supports the biological plausibility of linking TCE exposure and renal cancer.

What factors are associated with improvements in productivity in clinical laboratories in the Asia Pacific Region?

INTRODUCTION: Clinical laboratories usually have a quality management system such as ISO 15189, which provides a framework for quality and competence to perform medical testing and internal systems such as audit and nonconformance to ensure consistent processes. However, organizations need to have access to internal procedures and external competitors' performance to improve their operations. These are often seen as commercial or areas where it is difficult to agree on an acceptable goal. METHOD: In 2019, 1158 laboratories from 17 countries/regions in the Asia Pacific Region answered the survey, including 399 Chinese sites. The survey collected information on quality, turnaround time and productivity. RESULTS: Median productivity for laboratories in the Asia Pacific Region not including Chinese sites was 25 samples/FTE/day for small laboratories (workload: <250 samples/day), 100 for medium-sized laboratories (workload: 251-1000 samples/day) and 220 for large laboratories (workload: >1001 samples/day). The parameters associated with increased productivity in some laboratories were automation, middleware, Lean Six Sigma quality improvement activities and International Accreditation. CONCLUSION: This survey provides evidence of an association of quality improvement activities on laboratory productivity. There are differences in the effect of these activities in Chinese and non-Chinese laboratories in the Asia Pacific Region. The survey confirmed that the implementation of automation is associated with increased median productivity in all sites. Implementation of Lean Six Sigma and International Accreditation is associated with increased productivity in large laboratories.

Tetrahedral Framework Nucleic Acids Connected with MicroRNA-126 Mimics for Applications in Vascular Inflammation, Remodeling, and Homeostasis.

The repair of damaged endothelium is crucial for vascular homeostasis maintenance, which comprises the recovery of early stage impaired endothelial cells and migration of surrounding unimpaired endothelial cells. MicroRNAs (miRNAs) play an indispensable role in balancing gene expression in organisms. For vascular tissues, miR-126 is one of the most important regulators and might have substantial application potential in maintaining vascular homeostasis. In this study, a type of sticky-end-modified tetrahedral framework nucleic acids (tFNAs-SE) was employed to successfully link the miR-126 5p mimic duplex, which was termed tFNAs-miR-126 5p mimics (tFNAs-MMs). Existing vascular endothelial growth factors (VEGF), tFNAs-MMs can improve cell viability, resist apoptosis, and recover the state and functions of LPS-induced impaired human umbilical vein endothelial cells (HUVECs). The angiogenesis ability of impaired HUVECs was recovered by tFNAs-MMs in vitro and in vivo. The mechanisms underlying these phenomena were demonstrated to be related to the downregulation of caspase3 and negative regulators of VEGF (SPRED1 and PIK3R2). Moreover, tFNAs-MMs promoted the migration and proliferation of HUVECs. Briefly, the strategy of sticky-end-modified tFNAs connecting miRNA mimics is available for miRNA gain of function, while tFNAs-MMs might be a promising agent for repairing early stage vascular damage and maintaining vascular homeostasis.

Single cell transcriptomics reveals lineage trajectory of retinal ganglion cells in wild-type and Atoh7-null retinas.

Atoh7 has been believed to be essential for establishing the retinal ganglion cell (RGC) lineage, and Pou4f2 and Isl1 are known to regulate RGC specification and differentiation. Here we report our further study of the roles of these transcription factors. Using bulk RNA-seq, we identify genes regulated by the three transcription factors, which expand our understanding of the scope of downstream events. Using scRNA-seq on wild-type and mutant retinal cells, we reveal a transitional cell state of retinal progenitor cells (RPCs) co-marked by Atoh7 and other genes for different lineages and shared by all early retinal lineages. We further discover the unexpected emergence of the RGC lineage in the absence of Atoh7. We conclude that competence of RPCs for different retinal fates is defined by lineage-specific genes co-expressed in the transitional state and that Atoh7 defines the RGC competence and collaborates with other factors to shepherd transitional RPCs to the RGC lineage.

Occupational exposure to trichloroethylene is associated with a decline in lymphocyte subsets and soluble CD27 and CD30 markers.

Occupational cohort and case-control studies suggest that trichloroethylene (TCE) exposure may be associated with non-Hodgkin lymphoma (NHL) but findings are not consistent. There is a need for mechanistic studies to evaluate the biologic plausibility of this association. We carried out a cross-sectional molecular epidemiology study of 80 healthy workers that used TCE and 96 comparable unexposed controls in Guangdong, China. Personal exposure measurements were taken over a three-week period before blood collection. Ninety-six percent of workers were exposed to TCE below the current US Occupational Safety and Health Administration Permissible Exposure Limit (100 p.p.m. 8 h time-weighted average), with a mean (SD) of 22.2 (36.0) p.p.m. The total lymphocyte count and each of the major lymphocyte subsets including CD4+ T cells, CD8+ T cells, natural killer (NK) cells and B cells were significantly decreased among the TCE-exposed workers compared with controls (P < 0.05), with evidence of a dose-dependent decline. Further, there was a striking 61% decline in sCD27 plasma level and a 34% decline in sCD30 plasma level among TCE-exposed workers compared with controls. This is the first report that TCE exposure under the current Occupational Safety and Health Administration workplace standard is associated with a decline in all major lymphocyte subsets and sCD27 and sCD30, which play an important role in regulating cellular activity in subsets of T, B and NK cells and are associated with lymphocyte activation. Given that altered immunity is an established risk factor for NHL, these results add to the biologic plausibility that TCE is a possible lymphomagen.

Design, fabrication and applications of tetrahedral DNA nanostructure-based multifunctional complexes in drug delivery and biomedical treatment.

Although organic nanomaterials and inorganic nanoparticles possess inherent flexibility, facilitating functional modification, increased intracellular uptake and controllable drug release, their underlying cytotoxicity and lack of specificity still cause safety concerns. Owing to their merits, which include natural biocompatibility, structural stability, unsurpassed programmability, ease of internalization and editable functionality, tetrahedral DNA nanostructures show promising potential as an alternative vehicle for drug delivery and biomedical treatment. Here, we describe the design, fabrication, purification, characterization and potential biomedical applications of a self-assembling tetrahedral DNA nanostructure (TDN)-based multifunctional delivery system. First, relying on Watson-Crick base pairing, four single DNA strands form a simple and typical pyramid structure via one hybridization step. Then, the protocol details four different modification approaches, including replacing a short sequence of a single DNA strand by an antisense peptide nucleic acid, appending an aptamer to the vertex, direct incubation with small-molecular-weight drugs such as paclitaxel and wogonin and coating with protective agents such as cationic polymers. These modified TDN-based complexes promote the intracellular uptake and biostability of the delivered molecules, and show promise in the fields of targeted therapy, antibacterial and anticancer treatment and tissue regeneration. The entire duration of assembly and characterization depends on the cargo type and modification method, which takes from 2 h to 3 d.