Postnatal development of rat retina:a continuous observation and comparison between the organotypic retinal explant model and in vivo development

The organotypic retinal explant culture has been established for more than a decade and offers a range of unique advantages compared with in vivo experiments and cell cultures.However,the lack of systematic and continuous comparison between in vivo retinal development and the organotypic retinal explant culture makes this model controversial in postnatal retinal development studies.Thus,we aimed to verify the feasibility of using this model for postnatal retinal development studies by comparing it with the in vivo retina.In this study,we showed that postnatal retinal explants undergo normal development,and exhibit a consistent structure and timeline with retinas in vivo.Initially,we used SOX2 and PAX6 immunostaining to identify retinal progenitor cells.We then examined cell proliferation and migration by immunostaining with Ki-67 and doublecortin,respectively.Ki-67-and doublecortin-positive cells decreased in both in vivo and explants during postnatal retinogenesis,and exhibited a high degree of similarity in abundance and distribution between groups.Additionally,we used Ceh-10 homeodomain-containing homolog,glutamate-ammonia ligase(glutamine synthetase),neuronal nuclei,and ionized calcium-binding adapter molecule 1 immunostaining to examine the emergence of bipolar cells,Müller glia,mature neurons,and microglia,respectively.The timing and spatial patterns of the emergence of these cell types were remarkably consistent between in vivo and explant retinas.Our study showed that the organotypic retinal explant culture model had a high degree of consistency with the progression of in vivo early postnatal retina development.The findings confirm the accuracy and credibility of this model and support its use for long-term,systematic,and continuous observation.

3-photon fluorescence and third-harmonic generation imaging of myelin sheaths in mouse digital skin in vivo:A comparative study

Myelin sheaths wrapping axons are key structures that help maintain the propagation speed of action potentials in both central and peripheral nervous systems(CNS and PNS).However,noninvasive,deep imaging technologies visualizing myelin sheaths in the digital skin in vivo are lacking in animal models.3-photon°uorescence(3PF)imaging excited at the 1700-nm window enables deep imaging of myelin sheaths,but necessitates labeling by exogenous°uorescent dyes.Since myelin sheaths are lipid-rich structures which generate strong third-harmonic signals,in this paper,we perform a detailed comparative experimental study of both third-harmonic generation(THG)and 3PF imaging in the mouse digital skin in vivo.Our results show that THG imaging also enables visualization of myelin sheaths deep in the mouse digital skin,which shows colocalization with 3PF signals from labeled myelin sheaths.Besides its superior label-free advantage,THG does not su®er from photobleaching due to its 3PF property.

Deep-skin third-harmonic generation(THG)imaging in vivo excited at the 2200 nm window

The skin is heterogeneous and exerts strong scattering and aberration onto excitation light in multiphoton microscopy(MPM).Shifting to longer excitation wavelengths may help reduce skin scattering and aberration,potentially enabling larger imaging depths.However,previous demonstrations of skin MPM employ excitation wavelengths only up to the 1700 nm window,leaving an open question as to whether longer excitation wavelengths are suitable for deep-skin MPM.Here,in order to explore the longer-wavelength territory,first,we demonstrate characterization of the broadband transmittance of excised mouse skin,revealing a high transmittance window at 2200nm.Then,we demonstrate third-harmonic generation(THG)imaging in mouse skin in vivo excited at this window.With 9mW optical power on the skin surface operating at 1MHz repetition rate,we can get THG signals of 250
m below the skin surface.Comparative THG imaging excited at the 1700nm window shows that as imaging depth increases,THG signals decay even faster than those excited at 2200 nm.Our results thus uncover the 2200 nm window as a new,promising excitation window potential for deep-skin MPM.

Temporal and spatial distribution of metabotropic glutamate receptor 5 during development in the rat cortex and hippocampus

Metabotropic glutamate receptor 5 （mGluR5） is expressed by neurons in zones of active neurogenesis and is involved in the development of neural stem cells in vivo and in vitro. We examined the expression of mGluR5 in the cortex and hippocampus of rats during various prenatal and postnatal periods using immunohistochemistry. During prenatal development, mGluR5 was pdmadly localized to neuronal somas in the forebrain. During early postnatal periods, the receptor was mainly present on somas in the cortex, mGluR5 immunostaining was visible in apical dendrites and in the neuropil of neurons and persisted throughout postnatal development. During this period, pyramidal neurons were strongly labeled for the receptor. In the hippocampal CA1 region, mGluR5 immunoreactivity was more intense in the stratum oriens, stratum radiatum, and lacunosum moleculare at P0, P5 and P10 relative to P60. mGluR5 expression increased significantly in the molecular layer and decreased significantly in the granule cell layer of the dentate gyrus at P5, P10 and P60 in comparison with P0. Furthermore, some mGluR5-positive cells were also bromodeoxyuridine- or NeuroD-positive in the dentate gyrus at P14. These results demonstrate that mGluR5 has a differential expression pattern in the cortex and hippocampus during early growth, suggesting a role for this receptor in the control of domain specific brain developmental events.

The Anti-Proliferative Effect of 5-Fluorouracil on Tumor Is Highly Associated with the Renewal of Peripheral White Blood Cells

The efficacy of chemotherapy is thought to be direct killing of tumor cells, but documented studies have been shown that immunity plays a role in its effectiveness. In a pilot study to observe the bone marrow suppression and regeneration in tumor bearing mice induced by single dose injection of 5-fluorouracil (5-FU), we unexpectedly found that tumors grew fast as bone marrow mononuclear cells (BMC) and peripheral white blood cells (PWBC) were decreased quickly during myelosuppression meanwhile significantly slow as repopulating of BMC and PWBC during bone marrow regeneration after 5-FU treatment, no matter whether in low or high dose administration, but the higher the dose was, the lower of the nadir of BMC and PWBC were reached to, as well as the much more powerful duration and strength of the repopulated BMC and PWBC, suggested that the immunity might be a predominant drive in 5-FU chemotherapy. Due to the fact that BMC is the source of PWBC which is its final maturational and functional form, it could be proposed that the anti-proliferative effect of 5-FU on tumor is highly associated with the renewal of PWBC.

Optical trapping-enhanced probes designed by a deep learning approach

Realizing optical trapping enhancement is crucial in biomedicine,fundamental physics,and precision measurement.Taking the metamaterials with artificially engineered permittivity as photonic force probes in optical tweezers will offer unprecedented opportunities for optical trap enhancement.However,it usually involves multi-parameter optimization and requires lengthy calculations;thereby few studies remain despite decades of research on optical tweezers.Here,we introduce a deep learning(DL)model to attack this problem.The DL model can efficiently predict the maximum axial optical stiffness of Si∕Si_(3)N_(4)(SSN)multilayer metamaterial nanoparticles and reduce the design duration by about one order of magnitude.We experimentally demonstrate that the designed SSN nanoparticles show more than twofold and fivefold improvement in the lateral(k_(x)and k_(y))and the axial(k_(z))optical trap stiffness on the high refractive index amorphous TiO_(2)microsphere.Incorporating the DL model in optical manipulation systems will expedite the design and optimization processes,providing a means for developing various photonic force probes with specialized functional behaviors.

Identifying the potential transcriptional regulatory network in Hirschsprung disease by integrated analysis of microarray datasets

Objective Hirschsprung disease(HSCR)is one of the common neurocristopathies in children,which is associated with at least 20 genes and involves a complex regulatory mechanism.Transcriptional regulatory network(TRN)has been commonly reported in regulating gene expression and enteric nervous system development but remains to be investigated in HSCR.This study aimed to identify the potential TRN implicated in the pathogenesis and diagnosis of HSCR.Methods Based on three microarray datasets from the Gene Expression Omnibus database,the multiMiR package was used to investigate the microRNA(miRNA)-target interactions,followed by Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analyses.Then,we collected transcription factors(TFs)from the TransmiR database to construct the TF-miRNA-mRNA regulatory network and used cytoHubba to identify the key modules.Finally,the receiver operating characteristic(ROC)curve was determined and the integrated diagnostic models were established based on machine learning by the support vector machine method.Results We identified 58 hub differentially expressed microRNAs(DEMis)and 16 differentially expressed mRNAs(DEMs).The robust target genes of DEMis and DEMs mainly enriched in several GO/KEGG terms,including neurogenesis,cell-substrate adhesion,PI3K-Akt,Ras/mitogen-activated protein kinase and Rho/ROCK signaling.Moreover,2 TFs(TP53and TWIST1),4 miRNAs(has-miR-107,has-miR-10b-5p,has-miR-659-3p,and has-miR-371a-5p),and 4 mRNAs(PIM3,CHUK,F2RL1,and CA1)were identified to construct the TF-miRNA-mRNA regulatory network.ROC analysis revealed a strong diagnostic value of the key TRN regulons(all area under the curve values were more than 0.8).Conclusion This study suggests a potential role of the TF-miRNA-mRNA network that can help enrich the connotation of HSCR pathogenesis and diagnosis and provide new horizons for treatment.