A flexible electrode Array for genetic transfection of different layers of the retina by electroporation.

Electroporation (in which the permeability of a cell membrane is increased transiently by exposure to an appropriate electric field) has exhibited great potential of becoming an alternative to adeno-associated virus (AAV)-based retina gene delivery. Electroporation eliminates the safety concerns of employing exogenous viruses and exceeds the limit of AAV cargo size. Unfortunately, several concerns (e.g., relatively high electroporation voltage, poor surgical operability and a lack of spatial selectivity of retina tissue) have prevented electroporation from being approved for clinical application (or even clinical trials). In this study, a flexible micro-electrode array for retina electroporation (FERE) was developed for retina electroporation. A suitably shaped flexible substrate and well-placed micro-electrodes were designed to adapt to the retina curvature and generate an evenly distributed electric field on the retina with a significantly reduced electroporation voltage of 5 V. The FERE provided (for the first time) a capability of controlled gene delivery to the different structural layers of retina tissue by precise control of the distribution of the electrical field. After ensuring the surgical operability of the FERE on rabbit eyeballs, the FERE was verified to be capable of transfecting different layers of retina tissue with satisfactory efficiency and minimum damage. Our method bridges the technical gap between laboratory validation and clinical use of retina electroporation.

timsTOF HT Improves Protein Identification and Quantitative Reproducibility for Deep Unbiased Plasma Protein Biomarker Discovery.

Mass spectrometry (MS) is a valuable tool for plasma proteome profiling and disease biomarker discovery. However, wide-ranging plasma protein concentrations, along with technical and biological variabilities, present significant challenges for deep and reproducible protein quantitation. Here, we evaluated the qualitative and quantitative performance of timsTOF HT and timsTOF Pro 2 mass spectrometers for analysis of neat plasma samples (unfractionated) and plasma samples processed using the Proteograph Product Suite (Proteograph) that enables robust deep proteomics sampling prior to mass spectrometry. Samples were evaluated across a wide range of peptide loading masses and liquid chromatography (LC) gradients. We observed up to a 76% increase in total plasma peptide precursors identified and a >2-fold boost in quantifiable plasma peptide precursors (CV < 20%) with timsTOF HT compared to Pro 2. Additionally, approximately 4.5 fold more plasma peptide precursors were detected by both timsTOF HT and timsTOF Pro 2 in the Proteograph analyzed plasma vs neat plasma. In an exploratory analysis of 20 late-stage lung cancer and 20 control plasma samples with the Proteograph, which were expected to exhibit distinct proteomes, an approximate 50% increase in total and statistically significant plasma peptide precursors (q < 0.05) was observed with timsTOF HT compared to Pro 2. Our data demonstrate the superior performance of timsTOF HT for identifying and quantifying differences between biologically diverse samples, allowing for improved disease biomarker discovery in large cohort studies. Moreover, researchers can leverage data sets from this study to optimize their liquid chromatography-mass spectrometry (LC-MS) workflows for plasma protein profiling and biomarker discovery. (ProteomeXchange identifier: PXD047854 and PXD047839).

Human Pluripotent Stem Cells Derived Endothelial Cells Repair Choroidal Ischemia.

Choroidal atrophy is a common fundus pathological change closely related to the development of age-related macular degeneration (AMD), retinitis pigmentosa, and pathological myopia. Studies suggest that choroidal endothelial cells (CECs) that form the choriocapillaris vessels are the first cells lost in choroidal atrophy. It is found that endothelial cells derived from human pluripotent stem cells (hPSC-ECs) through the MESP1+ mesodermal progenitor stage express CECs-specific markers and can integrate into choriocapillaris. Single-cell RNA-seq (scRNA-seq) studies show that hPSC-ECs upregulate angiogenesis and immune-modulatory and neural protective genes after interacting with ex vivo ischemic choroid. In a rat model of choroidal ischemia (CI), transplantation of hPSC-ECs into the suprachoroidal space increases choroid thickness and vasculature density. Close-up examination shows that engrafted hPSC-ECs integrate with all layers of rat choroidal vessels and last 90 days. Remarkably, EC transplantation improves the visual function of CI rats. The work demonstrates that hPSC-ECs can be used to repair choroidal ischemia in the animal model, which may lead to a new therapy to alleviate choroidal atrophy implicated in dry AMD, pathological myopia, and other ocular diseases.

Highly Active and Water-Resistant Cu-Doped OMS-2 Catalysts for CO Oxidation: The Importance of the OMS-2 Synthesis Method and Cu Doping.

Porous cryptomelane-type Mn oxide (OMS-2) has an outstanding redox property, making it a highly desirable substitute for noble metal catalysts for CO oxidation, but its catalytic activity still needs to be improved, especially in the presence of water. Given the strong structure-performance correlation of OMS-2 for oxidation reactions, herein, OMS-2 is synthesized by solid state (OMS-2S), reflux (OMS-2R), and hydrothermal (OMS-2H) methods, aiming to improve its CO oxidation performance through manipulating synthesis parameters to tailor its particle size, morphology, and crystallinity. Characterization shows that OMS-2S has the highest CO oxidation activity in the absence of water due to its low crystallinity, high specific surface area, large oxygen vacancy content, and good redox property, but the presence of water can greatly reduce its CO oxidation activity. Doping Cu into an OMS-2 can not only improve its CO oxidation activity but also greatly improve its water tolerance. The Cu-doped OMS-2S catalyst with ∼4 wt % Cu can achieve a T90 of 49 °C (1% CO/10% O2/N2 and WHSV = 60,000 mL·g-1·h-1), ranking among the lowest reported T90 values for Mn oxide-based CO oxidation catalysts, and it can maintain nearly 100% CO conversion in the presence of 5 vol % water for over 50 h. In situ DRIFTs characterization indicates that the good water resistance of Cu-doped OMS-2S can be attributed to the significantly suppressed surface hydroxyl group generation because of Cu doping. This work demonstrates the importance of the synthesis method and Cu doping in determining the CO oxidation activity and water resistance of OMS-2 and will provide guidance for synthesizing highly active and water-resistant CO oxidation catalysts.

PKD1 participates in the processing of aortic dissection via regulating vascular smooth muscle cell contraction and MAPK signaling.

Aortic Dissection (AD) is a cardiovascular emergency with high mortality, of which one feature is the phenotypic switch of vascular smooth muscle cells (VSMCs). Transient Receptor Potential Channel Interacting (PKD1) has been regarded as one regulator as well as one biomarker for AD. However, multiple candidate pathways were reported though which PKD1 regulates AD in previous study, a comprehensive insight is still absent. In this study, we compared the AD and normal samples in transcriptome scale, and detected 717 PKD1-related differential expressed genes, which enriched in mitogen-activated protein kinase (MAPK) signal transduction (AD tissue preference) and VSMC contraction pathway (normal tissue preference). Furthermore, we also found two important functional hub genes in PKD1 regulation, JUN and ACTN2, and established a carnal-miRNA-mRNA network. Our study demonstrated the co-regulation of muscle development and signal transduction in AD's progression, and also provided the genetic basis for the following mechanism research with AD.

Conservation of beneficial microbes between the rhizosphere and the cyanosphere.

Biocrusts are phototroph-driven communities inhabiting arid soil surfaces. Like plants, most photoautotrophs (largely cyanobacteria) in biocrusts are thought to exchange fixed carbon for essential nutrients like nitrogen with cyanosphere bacteria. Here, we aim to compare beneficial interactions in rhizosphere and cyanosphere environments, including finding growth-promoting strains for hosts from both environments. To examine this, we performed a retrospective analysis of 16S rRNA gene sequencing datasets, host-microbe co-culture experiments between biocrust communities/biocrust isolates and a model grass (Brachypodium distachyon) or a dominant biocrust cyanobacterium (Microcoleus vaginatus), and metabolomic analysis. All 18 microbial phyla in the cyanosphere were also present in the rhizosphere, with additional 17 phyla uniquely found in the rhizosphere. The biocrust microbes promoted the growth of the model grass, and three biocrust isolates (Bosea sp._L1B56, Pseudarthrobacter sp._L1D14 and Pseudarthrobacter picheli_L1D33) significantly promoted the growth of both hosts. Moreover, pantothenic acid was produced by Pseudarthrobacter sp._L1D14 when grown on B. distachyon exudates, and supplementation of plant growth medium with this metabolite increased B. distachyon biomass by over 60%. These findings suggest that cyanobacteria and other diverse photoautotrophic hosts can be a source for new plant growth-promoting microbes and metabolites.

A Novel Surgical Approach for Big Sheet Allogenic Retinal Pigment Epithelium-Bruch Membrane Complex Transplantation Into the Subretinal Space.

PURPOSE: Allogenic transplantation of retinal pigmented epithelium monolayer sheet has experienced bottlenecks due to imperfect surgical techniques. In this study, we developed a novel approach for allogenic transplantation of big sheets of retinal pigment epithelium (RPE)-Bruch membrane complex. METHODS: RPE-Bruch membrane complex sheets of 5 × 6 mm2 to 10 × 10 mm2 were taken from donated eyes. Through a novel approach, the sheets of RPE-Bruch membrane complex were transplanted into the subretinal space of eight eyes (8 patients) with late-stage retinitis pigmentosa. The patients were followed up for 5 ± 2 months. RESULTS: All RPE-Bruch membrane complexes were successfully inserted into the subretinal space during the surgery. Follow-up examinations also showed that the grafts attached well to the transplantation site. No rejection or retinal detachment was found. CONCLUSION: Through our technique, big sheets of allogenic RPE-Bruch membrane complexes could be implanted into the subretinal space smoothly. This novel approach may be useful for big sheet of allogenic RPE-derived or stem cells-derived RPE transplantation in the treatment of RP and other retinal dystrophic diseases.

Ru-catalyzed C-H activation/cyclization of oximes with sulfoxonium ylides to access isoquinolines.

An external oxidant free Ru(II)-catalyzed C-H activation followed by an intermolecular annulation between oximes and sulfoxonium ylides has been developed. This transformation proceeds smoothly with a broad range of substrates, affording a series of isoquinoline derivatives in moderate to good yields. This protocol was successfully applied to the synthesis of moxaverine.

Zn deficiency disrupts Cu and S homeostasis in Chlamydomonas resulting in over accumulation of Cu and Cysteine.

Growth of Chlamydomonas reinhardtii in zinc (Zn) limited medium leads to disruption of copper (Cu) homeostasis, resulting in up to 40-fold Cu over-accumulation relative to its typical Cu quota. We show that Chlamydomonas controls its Cu quota by balancing Cu import and export, which is disrupted in a Zn deficient cell, thus establishing a mechanistic connection between Cu and Zn homeostasis. Transcriptomics, proteomics and elemental profiling revealed that Zn-limited Chlamydomonas cells up-regulate a subset of genes encoding "first responder" proteins involved in sulfur (S) assimilation and consequently accumulate more intracellular S, which is incorporated into L-cysteine, γ-glutamylcysteine, and homocysteine. Most prominently, in the absence of Zn, free L-cysteine is increased ∼80-fold, corresponding to ∼2.8 × 109 molecules/cell. Interestingly, classic S-containing metal binding ligands like glutathione and phytochelatins do not increase. X-ray fluorescence microscopy showed foci of S accumulation in Zn-limited cells that co-localize with Cu, phosphorus and calcium, consistent with Cu-thiol complexes in the acidocalcisome, the site of Cu(I) accumulation. Notably, cells that have been previously starved for Cu do not accumulate S or Cys, causally connecting cysteine synthesis with Cu accumulation. We suggest that cysteine is an in vivo Cu(I) ligand, perhaps ancestral, that buffers cytosolic Cu.

Hepatocyte-secreted FAM3D ameliorates hepatic steatosis by activating FPR1-hnRNP U-GR-SCAD pathway to enhance lipid oxidation.

Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disease worldwide; however, the underlying mechanisms remain poorly understood. FAM3D is a member of the FAM3 family; however, its role in hepatic glycolipid metabolism remains unknown. Serum FAM3D levels are positively correlated with fasting blood glucose levels in patients with diabetes. Hepatocytes express and secrete FAM3D, and its expression is increased in steatotic human and mouse livers. Hepatic FAM3D overexpression ameliorated hyperglycemia and steatosis in obese mice, whereas FAM3D-deficient mice exhibited exaggerated hyperglycemia and steatosis after high-fat diet (HFD)-feeding. In cultured hepatocytes, FAM3D overexpression or recombinant FAM3D protein (rFAM3D) treatment reduced gluconeogenesis and lipid deposition, which were blocked by anti-FAM3D antibodies or inhibition of its receptor, formyl peptide receptor 1 (FPR1). FPR1 overexpression suppressed gluconeogenesis and reduced lipid deposition in wild hepatocytes but not in FAM3D-deficient hepatocytes. The addition of rFAM3D restored FPR1's inhibitory effects on gluconeogenesis and lipid deposition in FAM3D-deficient hepatocytes. Hepatic FPR1 overexpression ameliorated hyperglycemia and steatosis in obese mice. RNA sequencing and DNA pull-down revealed that the FAM3D-FPR1 axis upregulated the expression of heterogeneous nuclear ribonucleoprotein U (hnRNP U), which recruits the glucocorticoid receptor (GR) to the promoter region of the short-chain acyl-CoA dehydrogenase (SCAD) gene, promoting its transcription to enhance lipid oxidation. Moreover, FAM3D-FPR1 axis also activates calmodulin-Akt pathway to suppress gluconeogenesis in hepatocytes. In conclusion, hepatocyte-secreted FAM3D activated the FPR1-hnRNP U-GR-SCAD pathway to enhance lipid oxidation in hepatocytes. Under obesity conditions, increased hepatic FAM3D expression is a compensatory mechanism against dysregulated glucose and lipid metabolism.

A novel predictive model for phthisis bulbi following facial hyaluronic acid cosmetic injection.

PURPOSE: To observe long-term prognosis of anterior segment ischemia (ASI) following hyaluronic acid (HA) injection, propose a severity grading system for ASI and a predictive model for phthisis bulbi (PB) based on long-term secretion dysfunction of ciliary process. METHODS: This is a retrospective case-control study. All enrolled 20 patients were divided into two groups and followed for at least 6 months to observe the formation and transformation characteristics of ASI and long-term prognosis based on the degrees of ciliary function damage. RESULTS: The severity of ASI following HA injection could be subdivided into 4 grades according to the degrees of ciliary function damage, comprising ASI grades 0, 1, 2 and 3. In 20 patients, ophthalmoplegia at 1-month follow-up, ASI within 1 month, ASI at 1-month follow-up, hypotony within 6 months were all significantly more common in study group than in control group (60% vs. 0%, P = 0.011; 100% vs. 20%, P = 0.001; 100% vs. 0%, P < 0.001; 80% vs. 0%, P = 0.001, respectively). Sensitivity, specificity and the area under the receiver operating characteristic curve (AUC) for predicting subsequent PB at 2-year follow-up through the co-occurrence of ophthalmoplegia at 1-month follow-up and hypotony within 6 months was 100%, 100% and 1.00, respectively. CONCLUSIONS: The new grading system for ASI and novel predictive model for PB we proposed could predict the long-term prognosis and probability of subsequent PB due to ASI following HA injection through several dynamic assessments within 6 months. LEVEL OF EVIDENCE: Level IV, observational prognostic study.

A modified aortic arch "island anastomosis" with stent graft technique for acute type A aortic dissection: a retrospective analysis.

BACKGROUND: A modified aortic arch "island anastomosis" with a stent graft technique was used in 33 patients with acute type A aortic dissection. We retrospectively reviewed our experience of this procedure and the short-term follow-up results. METHODS: This retrospective analysis included 33 patients with acute type A aortic dissection undergoing the modified aortic arch island anastomosis with stent graft procedure. Postoperatively, computed tomography angiography images were obtained before discharge and at 12 months. RESULTS: All patients underwent successful surgery without intraoperative death. Three patients received dialysis because of postoperative renal insufficiency, 1 patient received tracheotomy because of postoperative respiratory insufficiency, and 5 patients had postoperative delirium. Surgery caused stroke in 1 patient. No paraplegia was found, and no re-exploration for bleeding was performed. One patient died in the hospital due to multiple organ failure, and the other patients were discharged as expected. Only 1 patient had a proximal endoleak, and the patient was stable under close follow-up. The diameter of the descending thoracic aorta was smaller at 12 months postoperatively than preoperatively (34.5±2.5 mm versus 36.7±2.9 mm, P<0.05). The average diameter of the true lumen of the descending thoracic aorta was larger at 12 months postoperatively than preoperatively (24.1±3.1 mm versus 14.9±2.3 mm, P<0.05). CONCLUSIONS: The modified aortic arch island anastomosis with stent graft technique is a feasible and safety surgical strategy for acute type A aortic dissection. Short-term outcomes are satisfactory.

Cysteine: an ancestral Cu binding ligand in green algae?

Growth of Chlamydomonas reinhardtii in zinc (Zn) limited medium leads to disruption of copper (Cu) homeostasis, resulting in up to 40-fold Cu over-accumulation relative to its typical Cu quota. We show that Chlamydomonas controls its Cu quota by balancing Cu import and export, which is disrupted in a Zn deficient cell, thus establishing a mechanistic connection between Cu and Zn homeostasis. Transcriptomics, proteomics and elemental profiling revealed that Zn-limited Chlamydomonas cells up-regulate a subset of genes encoding "first responder" proteins involved in sulfur (S) assimilation and consequently accumulate more intracellular S, which is incorporated into L-cysteine, γ-glutamylcysteine and homocysteine. Most prominently, in the absence of Zn, free L-cysteine is increased ~80-fold, corresponding to ~ 2.8 × 10 9 molecules/cell. Interestingly, classic S-containing metal binding ligands like glutathione and phytochelatins do not increase. X-ray fluorescence microscopy showed foci of S accumulation in Zn-limited cells that co-localize with Cu, phosphorus and calcium, consistent with Cu-thiol complexes in the acidocalcisome, the site of Cu(I) accumulation. Notably, cells that have been previously starved for Cu do not accumulate S or Cys, causally connecting cysteine synthesis with Cu accumulation. We suggest that cysteine is an in vivo Cu(I) ligand, perhaps ancestral, that buffers cytosolic Cu.

Potential biomarkers of aortic dissection based on expression network analysis.

BACKGROUND: Aortic dissection (AD) is a rare disease with severe morbidity and high mortality. Presently, the pathogenesis of aortic dissection is still not completely clear, and studying its pathogenesis will have important clinical significance. METHODS: We downloaded 28 samples from the Gene Expression Omnibus (GEO) database (Accession numbers: GSE147026 and GSE190635), including 14 aortic dissection samples and 14 healthy controls (HC) samples. The Limma package was used to screen differentially expressed genes. The StarBasev2.0 tool was used to predict the upstream molecular circRNA of the selected miRNAs, and Cytoscape software was used to process the obtained data. STRING database was used to analyze the interacting protein pairs of differentially expressed genes under medium filtration conditions. The R package "org.hs.eg.db" was used for functional enrichment analysis. RESULTS: Two hundred genes associated with aortic dissection were screened. Functional enrichment analysis was performed based on these 200 genes. At the same time, 2720 paired miRNAs were predicted based on these 200 genes, among which hsa-miR-650, hsa-miR-625-5p, hsa-miR-491-5p and hsa-miR-760 paired mRNAs were the most. Based on these four miRNAs, 7106 pairs of circRNAs were predicted to be paired with them. The genes most related to these four miRNAs were screened from 200 differentially expressed genes (CDH2, AKT1, WNT5A, ADRB2, GNAI1, GNAI2, HGF, MCAM, DKK2, ISL1). CONCLUSIONS: The study demonstrates that miRNA-associated circRNA-mRNA networks are altered in AD, implying that miRNA may play a crucial role in regulating the onset and progression of AD. It may become a potential biomarker for the diagnosis and treatment of AD.

Radiomics analysis of contrast-enhanced T1W MRI: predicting the recurrence of acute pancreatitis.

To investigate the predictive value of radiomics based on T1-weighted contrast-enhanced MRI (CE-MRI) in forecasting the recurrence of acute pancreatitis (AP). A total of 201 patients with first-episode of acute pancreatitis were enrolled retrospectively (140 in the training cohort and 61 in the testing cohort), with 69 and 30 patients who experienced recurrence in each cohort, respectively. Quantitative image feature extraction was obtained from MR contrast-enhanced late arterial-phase images. The optimal radiomics features retained after dimensionality reduction were used to construct the radiomics model through logistic regression analysis, and the clinical characteristics were collected to construct the clinical model. The nomogram model was established by linearly integrating the clinically independent risk factor with the optimal radiomics signature. The five best radiomics features were determined by dimensionality reduction. The radiomics model had a higher area under the receiver operating characteristic curve (AUC) than the clinical model for estimating the recurrence of acute pancreatitis for both the training cohort (0.915 vs. 0.811, p = 0.020) and testing cohort (0.917 vs. 0.681, p = 0.002). The nomogram model showed good performance, with an AUC of 0.943 in the training cohort and 0.906 in the testing cohort. The radiomics model based on CE-MRI showed good performance for optimizing the individualized prediction of recurrent acute pancreatitis, which provides a reference for the prevention and treatment of recurrent pancreatitis.

Stomatin-like protein 2 deficiency exacerbates adverse cardiac remodeling.

Myocardial fibrosis, oxidative stress, and autophagy both play key roles in the progression of adverse cardiac remodeling. Stomatin-like protein 2 (SLP-2) is closely related to mitochondrial function, but little is known about its role and mechanism in cardiac remodeling. We developed doxorubicin (Dox), angiotensin (Ang) II, and myocardial ischemia-reperfusion (I/R) injury induced cardiac remodeling model and Dox treated H9C2 cell injury model using SLP-2 knockout (SLP-2-/-) mice and H9C2 cells with low SLP-2 expression. We first examined cardiac functional and structural changes as well as levels of oxidative stress, apoptosis and autophagy. We found that SLP-2 deficiency leads to decreased cardiac function and promotes myocardial fibrosis. After Dox and Ang II treatment, SLP-2 deficiency further aggravated myocardial fibrosis, increased myocardial oxidative stress and apoptosis, and activated autophagy by inhibiting PI3K-Akt-mTOR signaling pathway, ultimately exacerbating adverse cardiac remodeling. Similarly, SLP-2 deficiency further exacerbates adverse cardiac remodeling after myocardial I/R injury. Moreover, we extracted cardiomyocyte mitochondria for proteomic analysis, suggesting that SLP-2 deficiency may be involved in myocardial I/R injury induced adverse cardiac remodeling by influencing ubiquitination of intramitochondrial proteins. In addition, the oxidative stress, apoptosis and autophagy levels of H9C2 cells with low SLP-2 expression were further enhanced, and the PI3K-Akt-mTOR signaling pathway was further inhibited under Dox stimulation. Our results suggest that SLP-2 deficiency promotes myocardial fibrosis, disrupts normal mitochondrial function, overactivates autophagy via PI3K-Akt-mTOR signaling pathway, affects the level of ubiquitination, leads to irreversible myocardial damage, and ultimately exacerbates adverse cardiac remodeling.

Enhanced anaerobic hydrogen production from cotton straws assisted by copper molybdate.

Hydrogen production from dark fermentation has potential application due to its environmental friendliness, low production cost, and sustainability. However, there is still an obstacle to improving the efficiency of bioH2 production to meet the requirements in practical applications. In this research, copper molybdates are synthesized under different pH conditions as additives to study their different influence processes during anaerobic hydrogen production from cotton straws with the pure cultural system. A series of results indicate that CuMoO4 with appropriate experimental conditions has the highest H2 yield at 191.3 mL/g straws at 37 °C, which is 236% higher than the control group. It can be shown that O. ethanolica 8KG-4 has an obvious accompanying with high stability and low cytotoxicity for this clean energy production system as well as the improvement of metabolic pathway. These results extend new thinking of obtaining higher H2 yield as a biofuel in future production.

Development of a Surgical Technique for Subretinal Implants in Rats.

Retinal degeneration, such as age-related macular degeneration (AMD), is a leading cause of blindness worldwide. A myriad of approaches have been undertaken to develop regenerative medicine-based therapies for AMD, including stem cell-based therapies. Rodents as animal models for retinal degeneration are a foundation for translational research, due to the broad spectrum of strains that develop retinal degeneration diseases at different stages. However, mimicking human therapeutic delivery of subretinal implants in rodents is challenging, due to anatomical differences such as lens size and vitreous volume. This surgical protocol aims to provide a guided method for transplanting implants into the subretinal space in rats. A user-friendly comprehensive description of the critical steps has been included. This protocol has been developed as a cost-efficient surgical procedure for reproducibility across different preclinical studies in rats. Proper miniaturization of a human-sized implant is required prior to conducting the surgical experiment, which includes adjustments to the dimensions of the implant. An external approach is used instead of an intravitreal procedure to deliver the implant to the subretinal space. Using a small sharp needle, a scleral incision is performed in the temporal superior quadrant, followed by paracentesis to reduce intraocular pressure, thereby minimizing resistance during the surgical implantation. Next, a balanced salt solution (BSS) injection through the incision is carried out to achieve focal retinal detachment (RD). Lastly, insertion and visualization of the implant into the subretinal space are conducted. Post-operative assessment of the subretinal placement of the implant includes imaging by spectral domain optical coherence tomography (SD-OCT). Imaging follow-ups ascertain the subretinal stability of the implant, before the eyes are harvested and fixated for histological analysis.

Stomatin-like protein-2 attenuates macrophage pyroptosis and H9c2 cells apoptosis by protecting mitochondrial function.

Myocytes undergoing hypoxia condition can recruit macrophages and cause pro-inflammation initiation around the injury area. Mitochondrial dysfunction is related to macrophage pyroptosis. Stomatin-like protein-2 (SLP-2) can regulate mitochondrial biogenesis and function. Whether SLP-2 could affect macrophage pyroptosis remains unclear. In this study, bone marrow derived macrophages (BMDMs) were extracted from WT and SLP-2 knocked out mice, then stimulated by LPS/Nigericin. Western blot showed that SLP-2-/- promoted the expression of NLRP3, GSDMD-N, caspase-11 in macrophages, which means the deficiency of SLP-2 augments macrophage pyroptosis. Higher fluorescence intensity of dihydroethidium and TUNEL represented the increased ROS releasing and macrophage programmed death in SLP-2 deficiency groups. The immunofluorescence intensity of MtioTracker Red decreased and that of mitochondrial ROS (mtROS) increased in SLP-2 deletion groups with LPS/Nigericin stimulation, representing the increased mitochondrial damage. The expression level of HIF-1α increased in SLP-2 deletion macrophages with LPS and Nigericin stimulation. The level of Parkin and the ratio of LC3II/I decreased in SLP-2 deficiency macrophages after stimulated by LPS/Nigericin, compared with untreated macrophages. H9c2 cells were cultured in hypoxia condition before being cocultured with macrophage supernatant. The cocultured H9c2 cells were injured due to the serious pyroptosis of SLP-2 deficiency macrophages. According to these results, we suggest that SLP-2 can reduce macrophage pyroptosis and relieve hypoxia H9c2 cells injury through protecting mitochondrial function.

Overexpressed DDX3x promotes abdominal aortic aneurysm formation and activates AKT in ApoE knockout mice.

In recent years, abdominal aortic aneurysm (AAA) lesions have become one of the important diseases that threaten public health. Related studies have confirmed that the occurrence of abdominal aortic aneurysms is related to inflammatory stress, cell apoptosis, and elastic fiber degradation. DDX3x is thought to interact with inflammasomes such as NLRP3 to aggravate the process of the inflammatory response, but its role in the occurrence of AAA remains unclear. Since DDX3x is indispensable in animal embryonic growth, we used an adeno-associated virus to construct gene-overexpressing mice to induce aneurysm development through AngII infusion. The results indicated that the incidence of aneurysms, inflammatory cell infiltration, vascular smooth muscle cell transformation, and oxidative stress levels were significantly increased under the condition of DDX3x overexpression. At the signaling level, activation of the AKT pathway exacerbates aneurysm formation. Taken together, we believe that DDX3x plays a key role in the development of aneurysms and may be a new target for the treatment of aneurysm progression.