Berberine ameliorate inflammation and apoptosis via modulating PI3K/AKT/NFκB and MAPK pathway on dry eye.

BACKGROUND: Dry eye disease (DED) is a multifactorial disease in ocular surface, and inflammation plays an etiological role. Berberine (BBR) has shown efficacy in treating inflammatory diseases. Yet, there was no adequate information related to the therapeutic effects of BBR for DED. PURPOSE: To detect the effects and explore the potential mechanisms of BBR on DED. STUDY DESIGN: In vitro, in vivo study and network pharmacology analysis were involved. METHOD: The human corneal epithelium cells viability was evaluated with different concentrations of BBR. Dry eye murine model was established by exposing to the desiccating stress, and Ciclosporin (CSA), BBR eye drops or vehicle were topical administration for 7 days. The phenol red cotton tests, Oregon-green-dextran staining and Periodic acid-Schiff staining were performed and evaluated the dry eye after treatment. Inflammation and apoptosis levels of ocular surface were quantified. The potential targets related to berberine and dry eye were collected from databases. The Protein-Protein interaction network analysis and GO & KEGG enrichment analysis were realized by STRING database, Metascape platform and Cytoscape software to find core targets and signaling pathways. The SchrÖdinger software was used to molecular docking and PyMOL software to visualization. Finally, the levels of PI3K/AKT/NFκB and MAPK pathways were detected. RESULT: The data revealed BBR could rescue impaired HCE under hyperosmotic conditions. In addition, BBR eye drops could ameliorate dry eye. And BBR eye drops suppressed the inflammatory factors and CD4+T cells infiltration in conjunctiva. Besides, BBR eye drops protected ocular surface by avoiding the severe apoptosis and decreasing the level of MMP-3 and MMP-9. 148 common targets intersection between BBR and dry eye were found via network pharmacology analysis. Core proteins and core pathways were identified through PPI and GO&KEGG enrichment analysis. Molecular docking displayed excellent binding between BBR and those core targets. Finally, in vivo study verified that BBR eye drops had a therapeutic effect in dry eye by inhibiting PI3K/AKT/NFκB and MAPK pathways. CONCLUSION: The research provided convincing evidence that BBR could be a candidate drug for dry eye.

Ferroptosis drives photoreceptor degeneration in mice with defects in all-trans-retinal clearance.

The death of photoreceptor cells in dry age-related macular degeneration (AMD) and autosomal recessive Stargardt disease (STGD1) is closely associated with disruption in all-trans-retinal (atRAL) clearance in neural retina. In this study, we reveal that the overload of atRAL leads to photoreceptor degeneration through activating ferroptosis, a nonapoptotic form of cell death. Ferroptosis of photoreceptor cells induced by atRAL resulted from increased ferrous ion (Fe2+), elevated ACSL4 expression, system Xc- inhibition, and mitochondrial destruction. Fe2+ overload, tripeptide glutathione (GSH) depletion, and damaged mitochondria in photoreceptor cells exposed to atRAL provoked reactive oxygen species (ROS) production, which, together with ACSL4 activation, promoted lipid peroxidation and thereby evoked ferroptotic cell death. Moreover, exposure of photoreceptor cells to atRAL activated COX2, a well-accepted biomarker for ferroptosis onset. In addition to GSH supplement, inhibiting either Fe2+ by deferoxamine mesylate salt (DFO) or lipid peroxidation with ferrostatin-1 (Fer-1) protected photoreceptor cells from ferroptosis caused by atRAL. Abca4-/-Rdh8-/- mice exhibiting defects in atRAL clearance is an animal model for dry AMD and STGD1. We observed that ferroptosis was indeed present in neural retina of Abca4-/-Rdh8-/- mice after light exposure. More importantly, photoreceptor atrophy and ferroptosis in light-exposed Abca4-/-Rdh8-/- mice were effectively alleviated by intraperitoneally injected Fer-1, a selective inhibitor of ferroptosis. Our study suggests that ferroptosis is one of the important pathways of photoreceptor cell death in retinopathies arising from excess atRAL accumulation and should be pursued as a novel target for protection against dry AMD and STGD1.

Efficacy and Safety of Houttuynia Eye Drops Atomization Treatment for Meibomian Gland Dysfunction-Related Dry Eye Disease: A Randomized, Double-Blinded, Placebo-Controlled Clinical Trial.

PURPOSE: To evaluate the efficacy and safety of Houttuynia eye drops (a Chinese traditional medicine) atomization treatment in meibomian gland dysfunction (MGD)-related dry eye disease (DED) patients. METHODS: A total of 240 eligible patients diagnosed with MGD-related DED were assigned either Houttuynia eye drops or placebo for atomization once daily for four weeks in a multi-center, randomized, double-blind, placebo-controlled clinical study. Primary outcome evaluations used included eye symptom score (using the Chinese Dry Eye Questionnaire), meibum quality, and tear break-up time (TBUT), while safety evaluations included adverse events (AEs), visual acuity, and intraocular pressure monitoring. Indicators were measured at baseline as well as one week, two weeks, and four weeks after treatment. RESULTS: Primary outcome measures of the Houttuynia group were improved compared with their placebo counterparts following four-week treatment. Eye symptom scores were significantly reduced relative to the baseline in the Houttuynia group (mean ± standard error of the mean, 9.00 ± 0.61) compared with the placebo group (6.29 ± 0.55; p = 0.0018). Reduction in meibum quality score in the Houttuynia group (0.91 ± 0.10) was also significantly higher compared with the placebo group (0.57 ± 0.10; p = 0.0091), while TBUT in the treatment group (6.30 ± 0.22) was also longer than in the latter (5.60 ± 0.24; p = 0.0192). No medication-related adverse events were observed. CONCLUSIONS: Atomization treatment with Houttuynia eye drops is both clinically and statistically effective for the treatment of mild to moderate MGD-related DED patients. This approach is generally safe and was tolerated well by patients.

Multimodal Photoacoustic Imaging-Guided Regression of Corneal Neovascularization: A Non-Invasive and Safe Strategy.

Corneal neovascularization (CNV) is one of the main factors that induce blindness worldwide. However, current medical treatments cannot achieve non-invasive and safe inhibition of CNV. A noninvasive photoacoustic imaging (PAI)-guided method is purposed for the regression of CNV. PAI can monitor the oxygen saturation of cornea blood vessels through the endogenous contrast of hemoglobin and trace administrated drugs by themselves as exogenous contrast agents. An indocyanine green (ICG)-based nanocomposite (R-s-ICG) is prepared for CNV treatment via eye drops and subconjunctival injections. It is demonstrated that R-s-ICG can enrich corneal tissues and pathological blood vessels rapidly with minor residua in normal eyeball tissues. Anti-CNV treatment-driven changes in the blood vessels are assessed by real-time multimodal PAI in vivo, and then a safe laser irradiation strategy through the canthus is developed for phototherapy and gene therapy synergistic treatment. The treatment leads to the efficient inhibition of CNV with faint damages to normal tissues.

Onion Epithelial Membrane Scaffolds Transfer Corneal Epithelial Layers in Reconstruction Surgery.

Plants and their extracts have been used especially in China for more than ten centuries for preventing and treating disease. However, there are only few reports describing their use in animal cell culture and tissue transplantation. In this study, onion epithelial membranes (OEM) is used as scaffolds to support cultures of a variety of cells such as fibroblasts and epithelial cells notably; they maintain the phenotypic characteristics of corneal epithelial cells. This improvement includes preservation of the proliferative potential and stemness of rabbit corneal epithelial cells (RCECs). Such an outcome suggests that this cost-effective technology warrants further evaluation to determine if OEM is a viable candidate for use as scaffolds in corneal epithelial transplantation surgery. To test this possibility, rabbit corneal epithelial cells expanded on OEM are transplanted to treat corneal epithelial defects in limbal stem cell deficient rabbits. This procedure is successful because it shortens the time required for wound healing to restore losses in corneal epithelial integrity, and forms a more compact and stratified epithelium framework than the untreated group. Ultimately, should they be proven to be effective in other relevant animal model systems, their usefulness for treating wounds in a clinical setting warrants consideration.

All-trans-retinal dimer formation alleviates the cytotoxicity of all-trans-retinal in human retinal pigment epithelial cells.

Effective clearance of all-trans-retinal (atRAL) from retinal pigment epithelial (RPE) cells is important for avoiding its cytotoxicity. However, the metabolism of atRAL in RPE cells is poorly clarified. The present study was designed to analyze metabolic products of atRAL and to compare the cytotoxicity of atRAL versus its derivative all-trans-retinal dimer (atRAL-dimer) in human RPE cells. We found that all-trans-retinol (atROL) and a mixture of atRAL condensation metabolites including atRAL-dimer and A2E were generated after incubating RPE cells with atRAL for 6h, and the amount of atRAL-dimer was significantly higher than that of A2E. In the eyes of Rdh8-/- Abca4-/- mice, a mouse model with defects in retinoid cycle that displays some symbolic characteristics of age-related macular degeneration (AMD), the level of atRAL-dimer was increased compared to wild-type mice, and was even much greater than that of A2E & isomers. The cytotoxicity of atRAL-dimer was reduced compared with its precursor atRAL. The latter could provoke intracellular reactive oxygen species (ROS) overproduction, increase the mRNA expression of several oxidative stress related genes (Nrf2, HO-1, and γ-GCSh), and induce ΔΨm loss in RPE cells. By contrast, the abilities of atRAL-dimer to induce intracellular ROS and oxidative stress were much weaker versus that of concentration-matched atRAL, and atRAL-dimer exhibited no toxic effect on mitochondrial function at higher concentrations. In conclusion, the formation of atRAL-dimer during atRAL metabolic process ameliorates the cytotoxicity of atRAL by reducing oxidative stress.

Pain relief effect of breast feeding and music therapy during heel lance for healthy-term neonates in China: a randomized controlled trial.

OBJECTIVES: to test the effectiveness of breast feeding (BF), music therapy (MT), and combined breast feeding and music therapy (BF+MT) on pain relief in healthy-term neonates during heel lance. DESIGN: randomised controlled trial. SETTING: in the postpartum unit of one university-affiliated hospital in China from August 2013 to February 2014. PARTICIPANTS: among 288 healthy-term neonates recruited, 250 completed the trial. All neonates were undergoing heel lancing for metabolic screening, were breast fed, and had not been fed for the previous 30 minutes. INTERVENTIONS: all participants were randomly assigned into four groups - BF, MT, BF+MT, and no intervention - with 72 neonates in each group. Neonates in the control group received routine care. Neonates in the other three intervention groups received corresponding interventions five minutes before the heel lancing and throughout the whole procedure. MEASUREMENTS: Neonatal Infant Pain Scale (NIPS), latency to first cry, and duration of first crying. FINDINGS: mean changes in NIPS scores from baseline over time was dependent on the interventions given. Neonates in the BF and combined BF+MT groups had significantly longer latency to first cry, shorter duration of first crying, and lower pain mean score during and one minute after heel lance, compared to the other two groups. No significant difference in pain response was found between BF groups with or without music therapy. The MT group did not achieve a significantly reduced pain response in all outcome measures. CONCLUSIONS: BF could significantly reduce pain response in healthy-term neonates during heel lance. MT did not enhance the effect of pain relief of BF. IMPLICATIONS FOR PRACTICE: healthy-term neonates should be breast fed to alleviate pain during heel lance. There is no need for the additional input of classical music on breast feeding in clinic to relieve procedural pain. Nurses should encourage breast feeding to relieve pain during heel lance.

[The genes change of hMSC before and after the differentation into neuron-like cells].

OBJECTIVE: To study the genes change of hMSC before and after differentiation into neuron-like cells. METHODS: hMSC were separated from marrow, cultured and expanded in culture medium. After hMSC being induced to differentiate with Shenqiye, Neuron-specific enolase (MSE), neurofilament (NF), and glial fibrillary acidic protein (GFAP) were detected by immunocytochemistry respectively. Using semi-quantitiative RT-PCR to detect ten genes change in hMSC before and after differentiation. Two genes were used to detect the gene sequence. RESULTS: When treated with the inducer--Shenqiye, hMSC exhibited neuronal phenotype. The expressions of NSE and NF in the neuron-like cells were positive, but the glial astrocyte marker GFAP was not found. hMSC expressed germline, ectodermal, endodermal and mesodermal genes prior to neurogenasis. After differentiation, germline genes did not express, while the expressions of endodermal and mesodermal genes were weakened. Ectodermal and neuronal genes expressed stronger. CONCLUSION: hMSC express multipotentiality, and the neural differentiation comprises quantitative modulation of gene expression rather than simple on-off switching of neural specific genes.

[Experimental research on effect of human mesenchymal stem cells induced by shenqi fuzheng injection in cerebral infarction].

OBJECTIVE: To investigate the effect of shenqi fuzheng injection (SFI) in inducing differentiation of human mesenchymal stem cells (hMSCs) in brain stem and its effect on nervous function in model rats of cerebral infarction. METHODS: Middle cerebral artery occlusion model rats were made, and hMSCs was injected into their brain after being amplified in vitro and incubated with SFI for 0.5 h, then the survival, migration and differentiation of hMSCs in brain stem as well as the change of nervous function in model rats were observed. RESULTS: The post-transplantation reject reaction to hMSCs was low, it could survive as long as 6 weeks or more. No difference in area of infarction was shown before and after transplantation. Immunohistochemical staining showed that hMSCs expressed human neuron specific enolase (NSE), neurofilament (NF) and glial fibrillary acid protein (GFAP). The limb-kinetic function and tactile perception were improved in the model rats. CONCLUSION: SFI can induce hMSCs differentiate into neurons in vivo, and hMSCs may be the ideal germinal cells for treating cerebral infarction.

[Gene reversion of induced differentiation of adult human bone marrow-derived mesenchymal stem cells into neuron-like cells].

OBJECTIVE: To observe gene reversion during differentiation of human adult bone marrow-derived mesenchymal stem cells (MSCs) into neuron-like cells induced by Shenqiye. METHODS: The MSCs were separated, cultured and expanded in the culture medium and induced to differentiate into neuron-like cells with Shenqiye. The expressions of neuron-specific enolase (NSE), neurofilament (NF), and glial fibrillary acidic protein (GFAP) were detected by immunocytochemical method, and the changes of 10 genes of the cells after differentiation were detected by reverse transcriptional PCR. RESULTS: The MSCs exhibited neuronal phenotype when treated with Shenqiye and the neuron-like cells were positive for expressions of NSE and NF but not for glial astrocyte marker GFAP. After withdrawal of Shenqiye from the medium, the neuron-like cells were reversed to MSC, flat or spindal in morphology. The gene expression profiles of the redifferentiated cells were similar to those of undifferentiated MSCs. CONCLUSION: Shenqiye can induce MSC differentiation into neuron-like cells during and after which gene reversion may occur.