Bio-friendly long-term subcellular dynamic recording by self-supervised image enhancement microscopy.

Fluorescence microscopy has become an indispensable tool for revealing the dynamic regulation of cells and organelles. However, stochastic noise inherently restricts optical interrogation quality and exacerbates observation fidelity when balancing the joint demands of high frame rate, long-term recording and low phototoxicity. Here we propose DeepSeMi, a self-supervised-learning-based denoising framework capable of increasing signal-to-noise ratio by over 12 dB across various conditions. With the introduction of newly designed eccentric blind-spot convolution filters, DeepSeMi effectively denoises images with no loss of spatiotemporal resolution. In combination with confocal microscopy, DeepSeMi allows for recording organelle interactions in four colors at high frame rates across tens of thousands of frames, monitoring migrasomes and retractosomes over a half day, and imaging ultra-phototoxicity-sensitive Dictyostelium cells over thousands of frames. Through comprehensive validations across various samples and instruments, we prove DeepSeMi to be a versatile and biocompatible tool for breaking the shot-noise limit.

Endoscopic dacryocystorhinostomy with bicanalicular silicone tube intubation for treating chronic dacryocystitis secondary to nasolacrimal duct stent incarceration.

AIM: To investigate the feasibility of endoscopic dacryocystorhinostomy (En-DCR) with bicanalicular silicone tube intubation for treating chronic dacryocystitis secondary to nasolacrimal duct stent (NDS) incarceration. METHODS: En-DCRs were performed on 44 chronic dacryocystitis patients (46 eyes) secondary to NDS incarceration from April 2016 to October 2022. The granuloma and scar tissues were separated, and the removal of NDS incarceration was achieved during the surgery; the flap of the lacrimal sac was trimmed and anastomosed with nasal mucosal, a bicanalicular silicone tube was implanted, and lacrimal size and condition were assessed. The tube was removed 3mo after surgery. During the final follow-up of 12mo when the surgery was completed, the complications and the rates of surgical success were assessed. RESULTS: This study covered 40 patients (42 eyes). Intraoperatively, it was found that the lacrimal sac became small, and the sac wall had granulation and scar tissue attached to the incarcerated NDS in all eyes. At 12mo after surgery completed, the rates of the functional and anatomical success reached 80.95% (34/42) and 83.33% (35/42), respectively. Under the effect of intranasal ostial closure, seven eyes failed to achieve anatomical success. No serious complications (e.g., visual impairment, sinusitis, and orbital fat prolapse) was observed. CONCLUSION: With the success rate over 80% and no serious complications, En-DCR with bicanalicular silicone tube implantation is effective in treating chronic dacryocystitis secondary to NDS incarceration.

Comparison of Endoscopic Dacryocystorhinostomy Outcomes With or Without Bicanalicular Silicone Tube Intubation in Acquired Lacrimal Sac Mucocele.

PURPOSE: This paper was established to validate the necessity of bicanalicular silicone tube intubation (BSTI) in patients with acquired lacrimal sac mucocele (ALSM) receiving endoscopic dacryocystorhinostomy (DCR). METHODS: In total, 92 patients (92 eyes) diagnosed with ALSM undergone endoscopic DCR from November 2016 to December 2021 were recruited for our research. Patients were allocated into 2 equal treatment groups: group A (patients undergoing BSTI) and group B (patients not receiving this procedure). The tubes were removed 2 months postoperation in group A. Surgical outcomes and related complications were evaluated 12 months postoperation. RESULTS: Ultimately, this study included 83 patients with ALSM, consisting of 43 patients in group A and 40 patients in group B. All patients exhibited notable mass shrinkage with the opening of the lacrimal sac during the surgery and the swelling in the lacrimal sac area was completely relieved within 5 days postoperation. At the 12-month follow-up, no distinct difference was observed in the anatomic success rate between the 2 groups (group A: 93.0%; group B: 90.0%) ( t = 0.010, P > 0.05). However, patients in group A presented higher functional success rates (90.7%) in contrast to those patients in group B (72.5%) ( t = 4.635, P < 0.05). In both groups, the failure of the lacrimal passage reconstruction was attributed to granulation tissue formation or scar formation at the ostium. No sac mucocele recurrence occurred during the follow-up. CONCLUSION: Endoscopic DCR treatment for ALSM achieves satisfactory postoperative effects without recurrence, and BSTI may improve the functional success rate.

GSDME Increases Chemotherapeutic Drug Sensitivity by Inducing Pyroptosis in Retinoblastoma Cells.

Chemotherapy is an important part of retinoblastoma (RB) treatment. However, the development of drug resistance increases the likelihood of treatment failure. Therefore, increasing the sensitivity of chemotherapeutic drugs is very important. Recent research has explored the relationship between the expression level of gasdermin E (GSDME) and drug sensitivity in RB. Our study found that GSDME expression was significantly reduced in human RB tissues and cell lines. Downregulation of GSDME expression reduced the sensitivity of cells to chemotherapeutic drugs. Decitabine treatment and transfection with GSDME-overexpressing lentivirus (LV-GSDME) upregulated GSDME expression in Y79 and WERI-RB-1 cell lines. The half maximal inhibitory concentrations (IC50) for carboplatin-induced cell death were significantly reduced. Low-dose carboplatin could achieve the IC50, and no significant difference was found in the production of prodeath-activating proteins, but the mode of cell death changed from apoptosis to pyroptosis. Increased GSDME expression can reduce the required dose of chemotherapeutic drugs. After inhibition of caspase-3 activation, the IC50 of carboplatin-induced cell death was significantly increased in cells with high GSDME expression, and the method of cell death switched from pyroptosis to apoptosis, which increased the concentration of chemotherapeutic drugs. Furthermore, the sensitivity of cells to carboplatin was reduced. The in vivo xenograft tumor model further confirmed that GSDME upregulation could promote carboplatin-induced tumor cell death. Therefore, the sensitivity of cells to chemotherapeutic drugs can be predicted by detecting the GSDME expression level, and we used pyroptosis induction as a new method for promoting tumor death.

Clinical observation of ranibizumab combined with surgery in the treatment of neovascular glaucoma with vitreous hemorrhage.

PURPOSE: The aim was to explore the clinical efficacy of ranibizumab combined with surgical treatment of neovascular glaucoma with vitreous hemorrhage. MATERIALS AND METHODS: A total of 15 patients (17 affected eyes) who had neovascular glaucoma (NVG) with vitreous hemorrhage in our hospital were enrolled. After admission, the patient was given levofloxacin eye drops, 4 times a day. Three days later, the patients received intravitreal injection of ranibizumab. Then, trabeculectomy and vitrectomy were performed. The detailed clinical data, such as type of diseases, intraocular pressure (IOP), and best corrected visual acuity (BCVA), were collected before and after surgery. RESULTS: Visual acuity remained stable or improved in thirteen effected eyes and decreased in effected three eyes. Within 30 days after discharge, one effected eye recurred iris neovascularization with slightly higher IOP; then, the patient received intravitreal injection of ranibizumab again and neodymium-doped yttrium aluminum garnet (YAG) therapy. One patient (one effected eye) was given intravitreal ranibizumab injection again because of uncontrollable IOP and recurrence of neovascularization on iris surface and angle after operation; then, the patient received cyclophotocoagulation. Vitreous cavity hemorrhage occurred again in 3 patients after operation; then, these patients received the vitreous cavity lavage again. After trabeculectomy, inflammatory exudation or a small amount of bleeding could be seen in the anterior chamber of 6 young patients. CONCLUSION: Intravitreal injection of ranibizumab can effectively promote the rapid regression of intraocular neovascularization and help to control the IOP and improve postoperative visual acuity.

Mirror-enhanced scanning light-field microscopy for long-term high-speed 3D imaging with isotropic resolution.

Various biological behaviors can only be observed in 3D at high speed over the long term with low phototoxicity. Light-field microscopy (LFM) provides an elegant compact solution to record 3D information in a tomographic manner simultaneously, which can facilitate high photon efficiency. However, LFM still suffers from the missing-cone problem, leading to degraded axial resolution and ringing effects after deconvolution. Here, we propose a mirror-enhanced scanning LFM (MiSLFM) to achieve long-term high-speed 3D imaging at super-resolved axial resolution with a single objective, by fully exploiting the extended depth of field of LFM with a tilted mirror placed below samples. To establish the unique capabilities of MiSLFM, we performed extensive experiments, we observed various organelle interactions and intercellular interactions in different types of photosensitive cells under extremely low light conditions. Moreover, we demonstrated that superior axial resolution facilitates more robust blood cell tracking in zebrafish larvae at high speed.

Clinical characteristics, rates of blindness, and geographic features of PACD in China.

OBJECTIVE: To analyze the rates of blindness with the demographics and clinical characteristics of patients with primary angle-closure disease (PACD) to provide a comprehensive epidemiologic reference in China. METHODS: A retrospective analysis was conducted in the Chinese Glaucoma Study Consortium database, which is a national multicenter glaucoma research alliance of 111 hospitals participating between December 21, 2015 and September 9, 2018. The diagnosis of PACD was made by qualified physicians through examination. Comparison of sex, age, family history, subtypes of PACD, and blindness were analyzed. RESULTS: A total of 5762 glaucoma patients were included, of which 4588 (79.6%) had PACD. Of PACD patients, 72.1% were female with the sex ratio (F/M) of 2.6, and the average age of patients was 63.8±9.3 years with the majority between 60 and 70 years. Additionally, 30% of these patients had low vision in one eye, 8.8% had low vision in both eyes, 1.7% had blindness in one eye, and 0.3% had blindness in both eyes. There were statistical differences with regards to age between male and female patients with PACD, with male patients being older on average. Primary angle-closure glaucoma was more commonly diagnosed in males (60%) compared to females (35.9%), whereas acute primary angle closure (APAC) was more commonly diagnosed in females (54.3%) compared to males (37.7%). The visual acuity in APAC patients was lower and the rate of low vision and blindness was higher than other subtypes. CONCLUSION: PACD was the major type of glaucoma in Chinese hospitals. There were more female patients with PACD, mostly between 60 and 70 years old, with higher rates of APAC in women. APAC resulted in the worst visual outcomes of all PACD subtypes.

COPII mitigates ER stress by promoting formation of ER whorls.

Cells mitigate ER stress through the unfolded protein response (UPR). Here, we report formation of ER whorls as an effector mechanism of the ER stress response. We found that strong ER stress induces formation of ER whorls, which contain ER-resident proteins such as the Sec61 complex and PKR-like ER kinase (PERK). ER whorl formation is dependent on PERK kinase activity and is mediated by COPII machinery, which facilitates ER membrane budding to form tubular-vesicular ER whorl precursors. ER whorl precursors then go through Sec22b-mediated fusion to form ER whorls. We further show that ER whorls contribute to ER stress-induced translational inhibition by possibly modulating PERK activity and by sequestering translocons in a ribosome-free environment. We propose that formation of ER whorls reflects a new type of ER stress response that controls inhibition of protein translation.

Extraction and bioinformatics analysis of Chlamydia trachomatis LpxA.

OBJECTIVE: The aim of this study is to clone the LpxA gene of Chlamydia trachomatis and analyze its biological characteristics. METHODS: Specific primers were designed according to the sequence of Ct LpxA gene. LpxA gene was amplified by PCR and connected to pMD18-T vectors. Positive clones were selected for PCR and DNA sequencing. Finally, bioinformatics software was used to analyze the biological properties of LpxA protein. RESULTS: The total length of LpxA gene was 840 bp, encoding 280 amino acids. LpxA protein has no signal peptide and was located in bacterial cytoplasm. The prediction of secondary structure showed that the α-helix, extended strand, ß-turn and random coil accounted for 19.6%, 32.8%, 11.4% and 36%, respectively. According to the prediction of tertiary structure, three identical LpxA molecules constituted homologous trimers. It was predicted that there were 11 B cell epitopes in LpxA. CONCLUSION: Ct Lpxa gene was cloned, and LpxA protein structure and function were predicted.

Curcumin suppressed proliferation and migration of human retinoblastoma cells through modulating NF-κB pathway.

PURPOSE: To study the effect of curcumin on proliferation and invasion of the human retinoblastoma cells and its potential mechanism. METHODS: A cell line of retinoblastoma (WERI-Rb-1) was treated with various concentrations of curcumin (0-40 µM). Cell number was counted with CCK8 kit, and cell migration was assessed using the Transwell assay. Immunoblotting was performed to detect the proteins of metalloproteinase-2 (MMP-2), MMP-9 and vascular endothelial growth factor (VEGF) as well as nuclear translocation of nuclear factor-κB (NF-κB, p65). RESULTS: Proliferation and migration of WERI-Rb-1 cells were significantly inhibited by curcumin in a concentration-dependent manner (0-40 µM). Protein expressions of MMP-2, MMP-9 and VEGF in the WERI-Rb-1 cells were also significantly inhibited by curcumin in a concentration-dependent manner (0-40 µM). Furthermore, nuclear translocation of NF-κB (p65) was significantly inhibited by curcumin in time-dependent manner (6-24 h). CONCLUSION: Curcumin inhibited proliferation and migration of WERI-Rb-1 cells, a cell line of human retinoblastoma, which might be through modulating NF-κB and its downstream proteins including VEGF, MMP-2, and MMP-9.

The anti-malarial atovaquone selectively increases chemosensitivity in retinoblastoma via mitochondrial dysfunction-dependent oxidative damage and Akt/AMPK/mTOR inhibition.

Mitochondria has been identified as a promising target in several cancers. However, little is known on the effects of targeting mitochondria in retinoblastoma. In this work, we show that anti-malarial atovaquone, at clinically achievable concentration, demonstrates inhibitory effects to retinoblastoma cells, to a more extent than in normal retinal cells. Atovaquone also significantly increases chemosensitivity in retinoblastoma. Importantly, we show that retinoblastoma cells have higher level of mitochondrial respiration, membrane potential, mass and ATP compared to normal retinal cells. Although atovaquone significantly inhibits mitochondrial respiration and decrease ATP level in both malignant and normal retinal cells in a similar manner, atovaquone induces much more oxidative stress and damage in retinoblastoma than normal retinal cells. These suggest that normal retinal cells are more tolerable to mitochondrial dysfunctions than retinoblastoma cells. We further demonstrate that atovaquone targets Akt/AMPK/mTOR signaling via inducing mitochondrial dysfunction. Our pre-clinical work demonstrates the translational potential of atovaquone as an addition to the treatment armamentarium for retinoblastoma. Our work also demonstrates the differences of mitochondrial biogenesis and function in malignant versus normal retinal cells which are important for the targeted therapy in retinoblastoma.

Curcumin Alleviates Diabetic Retinopathy in Experimental Diabetic Rats.

PURPOSE: To investigate the potential protective effects of curcumin on the retina in diabetic rats. METHODS: An experimental diabetic rat model was induced by a low dose of streptozotocin combined with a high-energy diet. Rats which had blood glucose levels ≥11.6 mmol/L were used as diabetic rats. The diabetic rats were randomly divided into 3 groups: diabetic rats with no treatment (DM), diabetic rats treated with 100 mg/kg curcumin (DM + Cur 100 mg/kg), and diabetic rats treated with 200 mg/kg curcumin (DM + Cur 200 mg/kg). Curcumin was orally administered daily for 16 weeks. After 16 weeks of administration, the rats were euthanized, and eyes were dissected. Retinal histology was examined, and the thickness of the retina was measured. Ultrastructural changes of retinal ganglion cells, inner layer cells, retinal capillary, and membranous disks were observed by electron microscopy. Malondialdehyde, superoxide dismutase, and total antioxidant capacity were measured by ELISA. Expression levels of vascular endothelial growth factor (VEGF) in retina tissues were examined by immunohistochemical staining and ELISA. Expression levels of Bax and Bcl-2 in retina tissues were determined by immunohistochemical staining and Western blotting. RESULTS: Curcumin reduced the blood glucose levels of diabetic rats and decreased diabetes-induced body weight loss. Curcumin prevented attenuation of the retina in diabetic rats and ameliorated diabetes-induced ultrastructure changes of the retina, including thinning of the retina, apoptosis of the retinal ganglion cells and inner nuclear layer cells, thickening of retinal capillary basement membrane and disturbance of photoreceptor cell membranous disks. We also found that curcumin has a strong antioxidative ability in the retina of diabetic rats. It was observed that curcumin attenuated the expression of VEGF in the retina of diabetic rats. We also discovered that curcumin had an antiapoptotic effect by upregulating the expression of Bcl-2 and downregulating the expression of Bax in the retina of diabetic rats. CONCLUSIONS: Taken together, these results suggest that curcumin may have great therapeutic potential in the treatment of diabetic retinopathy which could be attributed to the hypoglycemic, antioxidant, VEGF-downregulating and neuroprotection properties of curcumin.