Optimal transcorneal electrical stimulation parameters for preserving photoreceptors in a mouse model of retinitis pigmentosa.

JOURNAL/nrgr/04.03/01300535-202419110-00034/figure1/v/2024-03-08T184507Z/r/image-tiff Retinitis pigmentosa is a hereditary retinal disease that affects rod and cone photoreceptors, leading to progressive photoreceptor loss. Previous research supports the beneficial effect of electrical stimulation on photoreceptor survival. This study aims to identify the most effective electrical stimulation parameters and functional advantages of transcorneal electrical stimulation (tcES) in mice affected by inherited retinal degeneration. Additionally, the study seeked to analyze the electric field that reaches the retina in both eyes in mice and post-mortem humans. In this study, we recorded waveforms and voltages directed to the retina during transcorneal electrical stimulation in C57BL/6J mice using an intraocular needle probe with rectangular, sine, and ramp waveforms. To investigate the functional effects of electrical stimulation on photoreceptors, we used human retinal explant cultures and rhodopsin knockout (Rho-/-) mice, demonstrating progressive photoreceptor degeneration with age. Human retinal explants isolated from the donors' eyes were then subjected to electrical stimulation and cultured for 48 hours to simulate the neurodegenerative environment in vitro. Photoreceptor density was evaluated by rhodopsin immunolabeling. In vivo Rho-/- mice were subjected to two 5-day series of daily transcorneal electrical stimulation using rectangular and ramp waveforms. Retinal function and visual perception of mice were evaluated by electroretinography and optomotor response (OMR), respectively. Immunolabeling was used to assess the morphological and biochemical changes of the photoreceptor and bipolar cells in mouse retinas. Oscilloscope recordings indicated effective delivery of rectangular, sine, and ramp waveforms to the retina by transcorneal electrical stimulation, of which the ramp waveform required the lowest voltage. Evaluation of the total conductive resistance of the post-mortem human compared to the mouse eyes indicated higher cornea-to-retina resistance in human eyes. The temperature recordings during and after electrical stimulation indicated no significant temperature change in vivo and only a subtle temperature increase in vitro (~0.5-1.5°C). Electrical stimulation increased photoreceptor survival in human retinal explant cultures, particularly at the ramp waveform. Transcorneal electrical stimulation (rectangular + ramp) waveforms significantly improved the survival and function of S and M-cones and enhanced visual acuity based on the optomotor response results. Histology and immunolabeling demonstrated increased photoreceptor survival, improved outer nuclear layer thickness, and increased bipolar cell sprouting in Rho-/- mice. These results indicate that transcorneal electrical stimulation effectively delivers the electrical field to the retina, improves photoreceptor survival in both human and mouse retinas, and increases visual function in Rho-/- mice. Combined rectangular and ramp waveform stimulation can promote photoreceptor survival in a minimally invasive fashion.

Characterizing the impact of species/strain-specific Lactiplantibacillus plantarum with community assembly and metabolic regulation in pickled Suancai.

To investigate the colonization and impact of the specific Lactiplantibacillus plantarum strains, four isolated strains were applied in pickled Suancai which is a traditional pickled mustard (Brassica juncea). Results showed that strain-8 with the highest lactic acid bacteria (LAB) counts and acetic acid (p < 0.05). There were 11.42 % ∼ 32.35 % differential volatile compounds detected, although nitriles, esters, and acids were predominant. L. plantarum disturbed the microbial community, in which the microbial composition of strain-11 was most similar to the naturally fermented sample. Amino acids, carbohydrate metabolism, and metabolism of cofactors and vitamins were the main functional classes because of the similar dominant microbes (Lactiplantibacillus and Levilactobacillus). The functional units were separated based on NMDS analysis, in which bacterial chemotaxis, amino acid-related units, biotin metabolism, fatty acid biosynthesis, and citrate cycle were significantly different calculated by metagenomeSeq and Benjamin-Hochberg methods (p < 0.05). The contents of most flavor compounds were consistent with their corresponding enzymes. In particular, glucosinolates metabolites were different and significantly related to the myrosinase and metabolic preference of LAB. Therefore, this study revealed the impact mechanism of the specific L. plantarum strains and provided a perspective for developing microbial resources to improve the flavor diversity of fermented vegetables.

Genomic DNA activates the AIM2 inflammasome and STING pathways to induce inflammation in lacrimal gland myoepithelial cells.

PURPOSE: Primary Sjögren's syndrome (pSS) is an autoimmune disease that mainly attacks the lacrimal glands causing severe aqueous-deficient dry eye. Clinical evidence indicates the DNA sensing mechanism in the pathogenesis of pSS. The purpose of the present study is to determine the pro-inflammatory effect of self-genomic DNA (gDNA) on myoepithelial cells (MECs), which along with acinar and ductal cells is a major cell type of the lacrimal gland. METHOD: MECs primary culture was acquired from female C57BL6J mice. Genomic DNA was extracted from the spleen of the same animal. The MECs were challenged with self-gDNA. The cytokine secretion was detected using supernatant by enzyme-linked immunosorbent assay (ELISA). The activation of inflammasomes was determined using FAM-FLICA. Cryosections of NOD.B10.H2b mouse model of pSS were obtained for immunofluorescence microscopy (IF), with Balb/C as control. RESULT: Treatment with gDNA activated AIM2 inflammasome assembly and function, leading to secretion of interleukin (IL)-1ß and IL-18 in MECs. The stimulation of IL-1ß secretion by gDNA appeared to be solely at the post-translational level, whereas IL-18 secretion was a combination of increased protein synthesis and post-translational modification. Genomic DNA also induced the activation of STimulators of INterferon Genes (STING), which correlated to the activation of STING in the lacrimal gland from the NOD.B10.H2b mouse. STING activation led to the secretion of IFN-ß via Nuclear Factor-κB (NF-κB). The IFN-ß further enhances the secretion of IL-1ß. The contractility of MECs was disabled by treatment with gDNA or poly AnT, independent of the level of intracellular [Ca2+]. CONCLUSION: Self-gDNA induces a proinflammatory response in lacrimal gland MECs by activating both the AIM2 inflammasome and STING and thus may contribute to the pathogenesis of pSS.

Purinergic 2X 4 (P2X4), but not P2X7, receptors increase cytosolic [Ca2+] and stimulate mucin secretion in rat conjunctival goblet cells to maintain ocular surface health.

Ionotropic purinergic receptors (P2XRs) are activated by ATP and ATP analogs. ATP can be released through ATP-permeable channels such as the pannexin hemichannels. Upon activation, the P2XRs become permeable to Ca2+, a potent stimulator of mucin secretion in conjunctival goblet cells (CGCs). The purpose of this study was to investigate the presence and function of P2XRs in CGCs. We also examined the presence of pannexin hemichannels. Rat first passage CGCs were stained with the goblet cell marker anti-cytokeratin 7 antibody and specific antibodies to P2X1-7 receptors and pannexin 1-3. mRNA expression was determined by RT-PCR using primers specific to P2XRs and pannexins. Proteins were identified with Western blotting (WB) using the same antibodies as for immunofluorescence (IF) microscopy. To study receptor function, CGCs were incubated with Fura 2-AM, exposed to agonists and antagonists, and intracellular [Ca2+] ([Ca2+]i) measured. [Ca2+]i was also measured after knock down of P2X4 and P2X7 receptor expression, and when exploiting P2XR specific characteristics. Lastly, mucin secretion was measured after the addition of several P2XR agonists. All P2XRs and pannexins were visualized with IF microscopy, and identified with RT-PCR and WB. [Ca2+]i was significantly increased when stimulated with ATP (10-7-10-4 M). Suramin, a non-selective P2XR antagonist at 10-4 M did not reduce ATP-induced peak [Ca2+]i. The potent P2X7 agonist, BzATP (10-7-10-4 M) increased the [Ca2+]i, although to a lesser extent than ATP. When measuring [Ca2+]i the effect of repeated applications of ATP at 10-5 or 10-6 M the response "desensitized" after 30-60 s. The P2X4 specific antagonist 5-BDBD decreased the P2X4 agonist, 2MeSATP,-induced [Ca2+]i increase. Furthermore, siRNA against the P2X4R, but not the P2X7R, decreased agonist-induced peak [Ca2+]i. ATP (10-5 M), BzATP (10-4 M) and 2MeSATP (10-5 M) induced mucin secretion. We conclude that all seven P2XRs are present in cultured rat CGCs. Of the P2XRs, only activation of the homotrimeric P2X4R appears to increase [Ca2+]i and induce mucin secretion. The P2X4R in CGCs offers a new therapeutic target for protective mucin secretion.

In Vitro Method to Study Sex-Based Differences in Conjunctival Goblet Cells.

Dry eye is a multi-factorial disease affecting ocular surface health, with a profoundly higher prevalence in women. Disruption of the gel-forming mucin that is secreted by conjunctival goblet cells (CGCs) onto the ocular surface contributes to multiple ocular surface diseases. The elimination of exogenous sex hormones is essential to obtain consistent results during in vitro study of sex-based differences in CGCs. This paper describes a method to minimize the presence of exogenous hormones in the study of sex-based differences in CGCs while maintaining their physiological function. CGCs from postmortem human donors of both sexes were cultured from pieces of the conjunctiva in RPMI medium with 10% fetal bovine serum (FBS) (referred to as the complete medium) until confluency. Nearly 48 h before the start of the experiments, CGCs were transferred to RPMI medium without phenol red or FBS but with 1% BSA (referred to as phenol-red-free medium). The normal cellular function was studied by measuring the increase in intracellular [Ca2+] ([Ca2+]i) after carbachol (Cch, 1 x 10-4 M) stimulation using fura 2/acetoxymethyl (AM) microscopy. The result shows that CGCs maintained normal function in the phenol-red-free media after 48 h. No significant difference in [Ca2+]i response was observed between phenol red-free RPMI medium and complete medium upon Cch stimulation. Therefore, we recommend using the phenol-red free RPMI medium with 1% BSA to eliminate exogenous hormones without altering the normal function of CGCs in the study of sex-based differences.

Anti-Inflammatory and Pro-Resolving Actions of the N-Terminal Peptides Ac2-26, Ac2-12, and Ac9-25 of Annexin A1 on Conjunctival Goblet Cell Function.

Annexin A1 (AnxA1) is the primary mediator of the anti-inflammatory actions of glucocorticoids. AnxA1 functions as a pro-resolving mediator in cultured rat conjunctival goblet cells to ensure tissue homeostasis through stimulation of intracellular [Ca2+] ([Ca2+]i) and mucin secretion. AnxA1 has several N-terminal peptides with anti-inflammatory properties of their own, including Ac2-26, Ac2-12, and Ac9-25. The increase in [Ca2+]i caused by AnxA1 and its N-terminal peptides in goblet cells was measured to determine the formyl peptide receptors used by the compounds and the action of the peptides on histamine stimulation. Changes in [Ca2+]i were determined by using a fluorescent Ca2+ indicator. AnxA1 and its peptides each activated formyl peptide receptors in goblet cells. AnxA1 and Ac2-26 at 10-12 mol/L and Ac2-12 at 10-9 mol/L inhibited the histamine-stimulated increase in [Ca2+]i, as did resolvin D1 and lipoxin A4 at 10-12 mol/L, whereas Ac9-25 did not. AnxA1 and Ac2-26 counter-regulated the H1 receptor through the p42/p44 mitogen-activated protein kinase/extracellular regulated kinase 1/2, ß-adrenergic receptor kinase, and protein kinase C pathways, whereas Ac2-12 counter-regulated only through ß-adrenergic receptor kinase. In conclusion, current data show that the N-terminal peptides Ac2-26 and Ac2-12, but not Ac9-25, share multiple functions with the full-length AnxA1 in goblet cells, including inhibition of histamine-stimulated increase in [Ca2+]i and counter-regulation of the H1 receptor. These actions suggest a potential pharmaceutical application of the AnxA1 N-terminal peptides Ac2-26 and Ac2-12 in homeostasis and ocular inflammatory diseases.

Effects of endogenous capsaicin stress and fermentation time on the microbial succession and flavor compounds of chili paste (a Chinese fermented chili pepper).

Chili paste, is a popular traditional product derived from chili pepper, and its fermentation system is affected by the variable concentration of capsaicin, which originates from the peppers. In the present study, the effects of capsaicin and fermentation time on the microbial community and flavor compounds of chili paste were investigated. After capsaicin supplementation, the total acid was significantly decreased (p < 0.05) along with lower total bacteria, especially lactic acid bacteria. Lactiplantibacillus, Lactobacillus, Weissella, Issatchenkia, Trichoderma, and Pichia were the shared and predominant genera; whereas, the Bacteroides and Kazachstania abundance was significantly increased due to the selection effect of capsaicin over time. Additionally, alterations of the microbial interaction networks and their metabolic preferences led to less lactic acid content with greater accumulation of ethyl nonanoate, methyl nonanoate, etc. This study will provide a perspective for selecting chili pepper varieties and improving the quality of fermented chili paste.

Lacrimal Gland Epithelial Cells Shape Immune Responses through the Modulation of Inflammasomes and Lipid Metabolism.

Lacrimal gland inflammation triggers dry eye disease through impaired tear secretion by the epithelium. As aberrant inflammasome activation occurs in autoimmune disorders including Sjögren's syndrome, we analyzed the inflammasome pathway during acute and chronic inflammation and investigated its potential regulators. Bacterial infection was mimicked by the intraglandular injection of lipopolysaccharide (LPS) and nigericin, known to activate the NLRP3 inflammasome. Acute injury of the lacrimal gland was induced by interleukin (IL)-1α injection. Chronic inflammation was studied using two Sjögren's syndrome models: diseased NOD.H2b compared to healthy BALBc mice and Thrombospondin-1-null (TSP-1-/-) compared to TSP-1WTC57BL/6J mice. Inflammasome activation was investigated by immunostaining using the R26ASC-citrine reporter mouse, by Western blotting, and by RNAseq. LPS/Nigericin, IL-1α and chronic inflammation induced inflammasomes in lacrimal gland epithelial cells. Acute and chronic inflammation of the lacrimal gland upregulated multiple inflammasome sensors, caspases 1/4, and interleukins Il1b and Il18. We also found increased IL-1ß maturation in Sjögren's syndrome models compared with healthy control lacrimal glands. Using RNA-seq data of regenerating lacrimal glands, we found that lipogenic genes were upregulated during the resolution of inflammation following acute injury. In chronically inflamed NOD.H2b lacrimal glands, an altered lipid metabolism was associated with disease progression: genes for cholesterol metabolism were upregulated, while genes involved in mitochondrial metabolism and fatty acid synthesis were downregulated, including peroxisome proliferator-activated receptor alpha (PPARα)/sterol regulatory element-binding 1 (SREBP-1)-dependent signaling. We conclude that epithelial cells can promote immune responses by forming inflammasomes, and that sustained inflammasome activation, together with an altered lipid metabolism, are key players of Sjögren's syndrome-like pathogenesis in the NOD.H2b mouse lacrimal gland by promoting epithelial dysfunction and inflammation.

Role of Oxidative Stress in Ocular Diseases: A Balancing Act.

Redox homeostasis is a delicate balancing act of maintaining appropriate levels of antioxidant defense mechanisms and reactive oxidizing oxygen and nitrogen species. Any disruption of this balance leads to oxidative stress, which is a key pathogenic factor in several ocular diseases. In this review, we present the current evidence for oxidative stress and mitochondrial dysfunction in conditions affecting both the anterior segment (e.g., dry eye disease, keratoconus, cataract) and posterior segment (age-related macular degeneration, proliferative vitreoretinopathy, diabetic retinopathy, glaucoma) of the human eye. We posit that further development of therapeutic interventions to promote pro-regenerative responses and maintenance of the redox balance may delay or prevent the progression of these major ocular pathologies. Continued efforts in this field will not only yield a better understanding of the molecular mechanisms underlying the pathogenesis of ocular diseases but also enable the identification of novel druggable redox targets and antioxidant therapies.

Transcorneal but not transpalpebral electrical stimulation disrupts mucin homeostasis of the ocular surface.

PURPOSE: Transcorneal electrical stimulation (TcES) is increasingly applied as a therapy for preserving and improving vision in retinal neurodegenerative and ischemic disorders. However, a common complaint about TcES is its induction of eye pain and dryness in the clinic, while the mechanisms remain unknown. METHOD: TcES or transpalpebral ES (TpES) was conducted in C57BL6j mice for 14 days. The contralateral eyes were used as non-stimulated controls. Levels of intracellular [Ca2+] ([Ca2+]i) were assessed by Fura-2AM. The conductance resistances of the eye under various ES conditions were measured in vivo by an oscilloscope. RESULTS: Although TcES did not affect tear production, it significantly induced damage to the ocular surface, as revealed by corneal fluorescein staining that was accompanied by significantly decreased mucin (MUC) 4 expression compared to the control. Similar effects of ES were detected in cultured primary corneal epithelium cells, showing decreased MUC4 and ZO-1 levels after the ES in vitro. In addition, TcES decreased secretion of MUC5AC from the conjunctiva in vivo, which was also corroborated in goblet cell cultures, where ES significantly attenuated carbachol-induced [Ca2+]i increase. In contrast to TcES, transpalpebral ES (TpES) did not induce corneal fluorescein staining while significantly increasing tear production. Importantly, the conductive resistance from orbital skin to the TpES was significantly smaller than that from the cornea to the retina in TcES. CONCLUSION: TcES, but not TpES, induces corneal epithelial damage in mice by disrupting mucin homeostasis. TpES thus may represent a safer and more effective ES approach for treating retinal neurodegeneration clinically.

Examining the relationship between academic stress and motivation toward physical education within a semester: A two-wave study with Chinese secondary school students.

The present study aimed to investigate the relationship between academic stress and motivation toward physical education (PE) through a longitudinal design with cross-lagged panel analyses. A sample of 556 Chinese secondary school students participated in the research and completed Perceived Locus of Causality Scale and Educational Stress Scale for Adolescents at the beginning of the semester and 3 months later. The results demonstrated that academic stress factors were positively related to less self-determined motivations except that worry about grades was positively related to more self-determined motivations within each time point. In addition, we found that academic stress negatively predicted more self-determined motivations but positively predicted less self-determined motivations, whereas worry about grades negatively predicted amotivation 3 months later. Meanwhile, the influence of amotivation on despondency was also found. These results suggest that academic stress can obstruct students' participation in PE through an impact on self-determined motivation. Our findings also indicate that self-determined students in PE will seek academic achievement as well, which in turn improves students' academic status.

Direct modulation of microglial function by electrical field.

Non-invasive electric stimulation (ES) employing a low-intensity electric current presents a potential therapeutic modality that can be applied for treating retinal and brain neurodegenerative disorders. As neurons are known to respond directly to ES, the effects of ES on glia cells are poorly studied. A key question is if ES directly mediates microglial function or modulates their activity merely via neuron-glial signaling. Here, we demonstrated the direct effects of ES on microglia in the BV-2 cells-an immortalized murine microglial cell line. The low current ES in a biphasic ramp waveform, but not that of rectangular or sine waveforms, significantly suppressed the motility and migration of BV-2 microglia in culture without causing cytotoxicity. This was associated with diminished cytoskeleton reorganization and microvilli formation in BV-2 cultures, as demonstrated by immunostaining of cytoskeletal proteins, F-actin and ß-tubulin, and scanning electron microscopy. Moreover, ES of a ramp waveform reduced microglial phagocytosis of fluorescent zymosan particles and suppressed lipopolysaccharide (LPS)-induced pro-inflammatory cytokine expression in BV-2 cells as shown by Proteome Profiler Mouse Cytokine Array. The results of quantitative PCR and immunostaining for cyclooxygenase-2, Interleukin 6, and Tumor Necrosis Factor-α corroborated the direct suppression of LPS-induced microglial responses by a ramp ES. Transcriptome profiling further demonstrated that ramp ES effectively suppressed nearly half of the LPS-induced genes, primarily relating to cellular motility, energy metabolism, and calcium signaling. Our results reveal a direct modulatory effect of ES on previously thought electrically "non-responsive" microglia and suggest a new avenue of employing ES for anti-inflammatory therapy.

Sex-based differences in conjunctival goblet cell responses to pro-inflammatory and pro-resolving mediators.

Many conjunctival inflammatory diseases differ between the sexes and altered conjunctival goblet cells (CGCs) response is often involved. Inflammation is initiated by the release of pro-inflammatory mediators and terminated by the biosynthesis of specialized pro-resolution mediators (SPMs). Herein, we determined the sex-based difference in the responses of CGCs to inflammatory stimuli or pro-resolving lipid SPMs and their interaction with sex hormones. GCs were cultured from pieces of human conjunctiva in RPMI media. CGCs were transferred 24 h before the start of experiments to phenol red-free and FBS-free media to minimize exogenous hormones. RT-PCR, immunofluorescence microscopy (IF), and Western Blot (WB) were performed to determine the presence of sex hormone receptors. Cellular response to pro-inflammatory stimuli or SPMs was studied by measuring the increase in intracellular [Ca2+] ([Ca2+]i) using fura 2/AM microscopy. Use of RT-PCR demonstrated estrogen receptor (ER) α in 4/5 males and 3/3 females; ERß in 2/4 males and 2/3 females; and androgen receptors (AR) in 3/3 male and 3/3 female CGCs. Positive immunoreactivity by IF and protein expression by WB was detected using antibodies for the ERα and ERß in 3/3 males and 3/3 females, while AR were only present in males. Significantly different Ca2+ responses between sexes were found with carbachol only at 10-3 M, but not with histamine or leukotriene (LT) B4 at any concentration used. Incubation with dihydrotestosterone (DHT), estrone (E1), or estradiol (E2) at 10-7 M for 30 min significantly inhibited the LTB4-stimulated [Ca2+]i increase in male and female CGCs. Incubation with DHT, E1, and E2 overnight significantly inhibited the LTB4 response in females, while DHT and E2 significantly inhibited the LTB4 response in males. The SPM lipoxin A4 (LXA4) (10-9-10-8 M), but not the resolvins D1 or D2, induced an [Ca2+]i increase that was significantly higher in males compared to females. We conclude that male and female CGCs showed differences in the expression of sex hormone receptors. Treatment with sex hormones altered pro-inflammatory mediator LTB4-induced response. Males compared to females have a higher response to the ω-6-fatty acid derived SPM LXA4, indicating males may terminate inflammation in conjunctival goblet cells faster than females.

Resolvin D2 uses multiple Ca2+ -dependent signaling pathways to stimulate mucin secretion in rat and human conjunctival goblet cells.

The mucin layer of the tear film is produced by goblet cells in the conjunctiva to protect the ocular surface and maintain homeostasis. The pro-resolving lipid mediator resolvin D2 (RvD2) biosynthesized from an omega 3 fatty acid actively terminates inflammation and regulates mucin secretion from conjunctival goblet cells. Our objective was to determine which Ca2+ -dependent signaling pathways RvD2 uses to stimulate conjunctival goblet cell function (CGC). We hypothesize that RvD2 activates multiple intracellular Ca2+ signaling pathways to stimulate CGC secretion. Rat and human CGCs were cultured from conjunctival explants. The amount of RvD2 receptor GPR18/DRV2 message and protein were determined. The intracellular concentration of Ca2+ ([Ca2+ ]i ) was measured in CGCs using a fluorescent Ca2+ dye and mucin secretion was determined by measuring protein secretion enzymatically with a lectin. Goblet cells were incubated with signaling pathway inhibitors before stimulation with RvD2 and [Ca2+ ]i or secretion was measured. In rat and human CGCs RvD2 receptor and in rat CGCs IP3 (a molecule that releases Ca2+ from intracellular organelles) receptors 1-3 were detected. In both species of CGC RvD2 increased [Ca2+ ]i similarly to RvD1. In rat CGCs, the increase in [Ca2+ ]i and secretion stimulated by RvD2 was significantly blocked by inhibitors to phospholipase (PL-) C and IP3 -receptor, but not protein kinase C. Increase in [Ca2+ ]i was blocked by the PLD inhibitor, but not the PLA2 inhibitor. Secretion was blocked by PLA2 inhibitor, but not the PLD inhibitor. An inhibitor of the epidermal growth factor receptor blocked the increase in [Ca2+ ]i by RvD2 in both species of CGCs. In CGCs RvD2 activates multiple intracellular signaling pathways that are Ca2+ -dependent, along with one Ca2+ -independent and one cAMP/protein kinase A-dependent pathway. Activation of these pathways stimulate mucin secretion from rat and human CGCs into the tear film contributing to ocular surface homeostasis and health.

Comparison of hyperreflective foci in macular edema secondary to multiple etiologies with spectral-domain optical coherence tomography: An observational study.

BACKGROUND: Hyperreflective foci (HRF) features in macular edema associated with different etiologies may indicate the disease pathogenesis and help to choose proper treatment. The goal of this study is to investigate the retinal microstructural features of macular edema (ME) secondary to multiple etiologies with spectral-domain optical coherence tomography (SD-OCT) and analyze the origin of HRF in ME. METHODS: This was a retrospective study. SD-OCT images were reviewed to investigate macular microstructural features such as the number and distribution of HRF and hard exudates and the internal reflectivity of the cysts. The differences in microstructural features between groups and the correlations between the number of HRF and other parameters were analyzed. RESULTS: A total of 101 eyes with ME from 86 diabetic (diabetic macular edema, DME) patients, 51 eyes from 51 patients with ME secondary to branch retinal vein occlusion (branch retinal vein occlusion-macular edema, BRVO-ME), 59 eyes from 58 central retinal vein occlusion (central retinal vein occlusion-macular edema, CRVO-ME) patients, and 26 eyes from 22 uveitis (uveitic macular edema, UME) patients were included in this study. The number of HRF, the frequency of hard exudates and the enhanced internal reflectivity of the cysts were significantly different among the groups. The number of HRF in the DME group was significantly higher than that in the other groups (all P < 0.05). The frequency of hard exudates and enhanced internal reflectivity of the cysts in the DME group were significantly higher than ME secondary to other etiologies (all P < 0.001). Within the DME group, the number of HRF in the patients with hard exudates was significantly higher than that in the patients without hard exudates (P < 0.001). CONCLUSION: HRF detected with SD-OCT were more frequent in DME patients than in BRVO-ME, CRVO-ME, or UME patients. The occurrence of HRF was correlated with the frequency of hard exudates. HRF may result from the deposition of macromolecular exudates in the retina, which is speculated to be a precursor of hard exudates.

Fucosylation Promotes Cytolytic Function and Accumulation of NK Cells in B Cell Lymphoma.

Natural killer (NK) cells have been demonstrated as a promising cellular therapy as they exert potent anti-tumor immune responses. However, applications of NK cells to tumor immunotherapy, especially in the treatment of advanced hematopoietic and solid malignancies, are still limited due to the compromised survival and short persistence of the transferred NK cells in vivo. Here, we observed that fucosyltransferase (FUT) 7 and 8 were highly expressed on NK cells, and the expression of CLA was positively correlated with the accumulation of NK cells in clinical B cell lymphoma development. Via enzyme-mediated ex vivo cell-surface fucosylation, the cytolytic effect of NK cells against B cell lymphoma was significantly augmented. Fucosylation also promoted NK cell accumulation in B cell lymphoma-targeted tissues by enhancing their binding to E-selectin. Moreover, fucosylation of NK cells also facilitated stronger T cell anti-tumor immune responses. These findings suggest that ex vivo fucosylation contributes to enhancing the effector functions of NK cells and may serve as a novel strategy for tumor immunotherapy.

Baicalein-A Potent Pro-Homeostatic Regulator of Microglia in Retinal Ischemic Injury.

Retinal ischemia is a common cause of many retinal diseases, leading to irreversible vision impairment and blindness. Excessive neuroinflammation, including microglial activation and T-cell responses, has been identified as a critical factor associated with neurodegeneration in retinal ischemia. Baicalein is a natural flavonoid reported to have broad anti-inflammatory and neuroprotective bioactivities. Herein, the effects of baicalein on microglia activation in vitro and in vivo were investigated. We found that baicalein exhibited robust anti-inflammatory effect on cultured human and mouse microglia, as demonstrated by decreased induction of pro-inflammatory cytokines and the phosphorylation of phosphoinositide 3-kinase (PI3K) and nuclear factor kappa B (NFκB). Proteomic analysis further unraveled baicalein's effect on modulating IL-17 signaling pathways and its upstream regulator IL-1ß. Intravitreal administration of baicalein in the mouse model of retinal ischemia/reperfusion (I/R) injury attenuated microglial activation and retinal T-cell infiltration, particularly the T helper 17 cells. Additionally, baicalein was shown to exert neuroprotective effects by significantly reducing the retinal ganglion cell (RGC) loss after I/R injury, leading to an improved retinal function and spatial vision. These results suggest that baicalein, a natural flavonoid, acts as a negative regulator of activated microglia and immune responses both in vitro and in vivo, effectively alleviating neurodegeneration in retinal I/R injury. This finding indicates that baicalein could be a potential therapeutic agent against currently incurable degenerative retinal diseases.

Exploring major variable factors influencing flavor and microbial characteristics of Pixian Doubanjiang.

Pixian Doubanjiang (PXDBJ) is a typical Asian condiment with a unique spicy flavor produced by traditional spontaneous solid-state fermentation. In this study, flavor and the microbial community characteristics of PXDBJ were investigated based on polyphasic detection approaches, and further revealed the potential regulations of the variable factors on its characteristics. Specifically, the chili varieties and the ripening periods affected the features of organic acids and amino acids, respectively. PLS-DA plot demonstrated that the volatiles contents varied by the intermediate product Banpei and the ripening periods although the predominant aroma compounds were roughly the same. Those results were verified by Spearman's arithmetic (P < 0.05), and established a chili variety-classification model based on OPLS-DA plot (Q2(cum) = 0.918, P < 0.05). In addition, the relative abundance of Saccharomycetales, Zygosaccharomyces, and Muribaculaceae varied with ripening periods, and further revealed the contributions of those variable factors to the flavor and correlated with the predominant microbes. This comprehensive study on the raw materials and spatiotemporal characteristics is valuable for exploring the fermentation mechanism and controlling the product quality.

The effects of different coculture patterns with salt-tolerant yeast strains on the microbial community and metabolites of soy sauce moromi.

In this study, the contributions of three strains and different coculture patterns to microbial community diversity and metabolites in the high-salt liquid-state fermentation (HLF) soy sauce moromi were investigated. A comparison of two strains of Zygosaccharomyces rouxii showed that strain QH-25 had a stronger ability to contribute metabolites, including both nonvolatile and volatile types, to the moromi than strain QH-1, except for volatile acids and ketones. Of the various fortification patterns tested, the content of metabolites was significantly increased by inoculating Z. rouxii QH-25 prior to C. versatilis, especially the content of volatiles, including ketones, esters, phenols, and alcohols, which increased 1.61-fold compared with those in the control sample; the contents of these components were increased 3.07-, 1.91-, 1.36-, and 1.22-fold, respectively. In particular, characteristic components such as ethyl octanoate, 4-hydroxy-2(or 5)-ethyl-5(or 2)-methyl-3(2H)-furanone (HEMF), 4-ethyl-2-methoxy-phenol (4-EG), and 3-methyl-1-butanol were increased 3.99-, 3.29-, 1.63-, and 0.70-fold, respectively. Redundancy analysis (RDA) showed that Staphylococcus, Zygosaccharomyces, and Candida were positively correlated with the unique components described above. In addition, the nodes of the interaction network between Zygosaccharomyces and Candida were increased, and the positive correlation of Zygosaccharomyces with Staphylococcus was enhanced by inoculating Z. rouxii prior to C. versatilis. These results suggested that the unique flavor of soy sauce was closely related to the metabolic characteristics of strains affiliated with Z. rouxii, whether cultured singly or cocultured with C. versatilis. This study also provided a reference method for determining the differences in community structure and metabolites between traditional techniques and modern processes for soy sauce fermentation.

The Prognostic Value of Early Detection of Minimal Residual Disease as Defined by Flow Cytometry and Gene Mutation Clearance for Myelodysplastic Syndrome Patients After Myeloablative Allogeneic Hematopoietic Stem-Cell Transplantation.

High relapse incidence remains a major problem for myelodysplastic syndrome (MDS) patients who have received an allogeneic hematopoietic stem-cell transplantation (allo-HSCT). We retrospectively analyzed the correlations between clinical outcomes and minimal residual disease (MRD) by using mutations (MUT) and flow cytometry (FCM) analysis of 115 MDS patients with allo-HSCT. We divided 115 MDS patients into four groups based on molecular genetics and FCM MRD results at day 30 post-HSCT. There were significant differences in the 2-year progression-free survival (PFS) between the FCMhigh MUTpos and FCMlow MUTneg groups (20% vs 79%, P < 0.001). In addition, by univariate analysis, we found that an IPSS-R score ≥4 pre-HSCT (HR, 5.061; P=0.007), DNMT3A mutations (HR, 2.291; P=0.052), TP53 mutations (HR, 3.946; P=0.011), and poor and very poor revised International Prognostic Scoring System (IPSS-R) cytogenetic risk (HR, 4.906; P < 0.001) were poor risk factors for PFS. In multivariate analysis, we found that an IPSS-R score ≥ 4 pre-HSCT (HR, 4.488; P=0.015), DNMT3A mutations (HR, 2.385; P=0.049), positive FCM MRD combined with persistence gene mutations at day 30 (HR, 5.198; P=0.013) were independent risk factors for disease progression. In conclusion, our data indicated that monitoring MRD by FCM combined with gene mutation clearance at day 30 could help in the prediction of disease progression for MDS patients after transplantation.