TRIM25 inhibition attenuates inflammation, senescence, and oxidative stress in microvascular endothelial cells induced by hyperglycemia.

PURPOSES: This work aimed to assess the possible role of TRIM25 in regulating hyperglycemia-induced inflammation, senescence, and oxidative stress in retinal microvascular endothelial cells, all of which exert critical roles in the pathological process of diabetic retinopathy. METHODS: The effects of TRIM25 were investigated using streptozotocin-induced diabetic mice, human primary retinal microvascular endothelial cells cultured in high glucose, and adenoviruses for TRIM25 knockdown and overexpression. TRIM25 expression was evaluated by western blot and immunofluorescence staining. Inflammatory cytokines were detected by western blot and quantitative real-time PCR. Cellular senescence level was assessed by detecting senescent marker p21 and senescence-associated-ß-galactosidase activity. The oxidative stress state was accessed by detecting reactive oxygen species and mitochondrial superoxide dismutase. RESULTS: TRIM25 expression is elevated in the endothelial cells of the retinal fibrovascular membrane from diabetic patients compared with that of the macular epiretinal membrane from non-diabetic patients. Moreover, we have also observed a significant increase in TRIM25 expression in diabetic mouse retina and retinal microvascular endothelial cells under hyperglycemia. TRIM25 knockdown suppressed hyperglycemia-induced inflammation, senescence, and oxidative stress in human primary retinal microvascular endothelial cells while TRIM25 overexpression further aggregates those injuries. Further investigation revealed that TRIM25 promoted the inflammatory responses mediated by the TNF-α/NF-κB pathway and TRIM25 knockdown improved cellular senescence by increasing SIRT3. However, TRIM25 knockdown alleviated the oxidative stress independent of both SIRT3 and mitochondrial biogenesis. CONCLUSION: Our study proposed TRIM25 as a potential therapeutic target for the protection of microvascular function during the progression of diabetic retinopathy.

Quantitative evaluation of ocular vascularity and correlation analysis in patients with diabetic retinopathy by SMI and OCTA.

AIMS: To find potential relation between retrobulbar vessels and fundus microvessels and to detect sensitive and effective clinical indicators in predicting the progress of diabetic retinopathy (DR), ocular hemodynamics were measured using superb microvascular imaging (SMI) and ultrawide-field optical coherence tomography angiography (UWF-OCTA). METHODS: Observational, cross-sectional study evaluating ocular hemodynamics in patients with DR by SMI (Aplio i900, Canon Medical) and UWF-OCTA (BM-400 K BMizar, Tupai Medical Technology). The peak systolic velocity (PSV), end-diastolic velocity (EDV), and resistive index (RI) of the central retinal artery (CRA), posterior ciliary artery (PCA), and ophthalmic artery (OA) were measured by SMI. UWF-OCTA evaluated the fundus vascular parameters. A correlation analysis was used to determine the correlation between SMI and UWF-OCTA parameters. RESULTS: One hundred thirty-nine eyes of 139 diabetic patients were included: 29 without DR (NDR), 36 with mild to moderate nonproliferative DR (M-NPDR), 37 with severe NPDR (S-NPDR), and 37 with proliferative DR (PDR). PSV and EDV of retrobulbar vessels decreased from NDR to S-NPDR while increasing PDR. RI of OA showed a decreasing trend in the progression of DR, but other vessels didn't show the same trend. ROC curve analysis showed that CRAPSV, CRAEDV, PCAEDV, OAPSV, and OAEDV had diagnostic value distinguishing M-NPDR and S-NPDR. The correlation analysis observed a significant association between the SMI parameters of CRA and PCA and UWF-OCTA parameters. CRA hemodynamics were more associated with fundus vascular parameters, especially the retina, in the NDR group than in the M-NPDR group. In contrast, PCA consistently correlated with fundus vascular parameters, especially in the choroid, from the NDR to the M-NPDR group. However, OA showed a poor correlation with OCTA parameters. CONCLUSION: The velocity of retrobulbar vessels, mainly the CRA, may serve as a valuable predictor for assessing the progress of DR. The use of SMI in diabetic patients may help identify patients at risk of developing retinopathy.

Integration of Vitreous Lipidomics and Metabolomics for Comprehensive Understanding of the Pathogenesis of Proliferative Diabetic Retinopathy.

As a vision-threatening complication of diabetes mellitus (DM), proliferative diabetic retinopathy (PDR) is associated with sustained metabolic disorders. Herein, we collected the vitreous cavity fluid of 49 patients with PDR and 23 control subjects without DM for metabolomics and lipidomics analyses. Multivariate statistical methods were performed to explore relationships between samples. For each group of metabolites, gene set variation analysis scores were generated, and we constructed a lipid network by using weighted gene co-expression network analysis. The association between lipid co-expression modules and metabolite set scores was investigated using the two-way orthogonal partial least squares (O2PLS) model. A total of 390 lipids and 314 metabolites were identified. Multivariate statistical analysis revealed significant vitreous metabolic and lipid differences between PDR and controls. Pathway analysis showed that 8 metabolic processes might be associated with the development of PDR, and 14 lipid species were found to be altered in PDR patients. Combining metabolomics and lipidomics, we identified fatty acid desaturase 2 (FADS2) as an important potential contributor to the pathogenesis of PDR. Collectively, this study integrates vitreous metabolomics and lipidomics to comprehensively unravel metabolic dysregulation and identifies genetic variants associated with altered lipid species in the mechanistic pathways for PDR.

The single-cell landscape of alternative transcription start sites of diabetic retina.

More than half of mammalian protein-coding genes have multiple transcription start sites. Alternative transcription start site (TSS) modulate mRNA stability, localization, and translation efficiency on post-transcription level, and even generate novel protein isoforms. However, differential TSS usage among cell types in healthy and diabetic retina remains poorly characterized. In this study, by using 5'-tag-based single-cell RNA sequencing, we identified cell type-specific alternative TSS events and key transcription factors for each of retinal cell types. We observed that lengthening of 5'- UTRs in retinal cell types are enriched for multiple RNA binding protein binding sites, including splicing regulators Rbfox1/2/3 and Nova1. Furthermore, by comparing TSS expression between healthy and diabetic retina, we identified elevated apoptosis signal in Müller glia and microglia, which can be served as a putative early sign of diabetic retinopathy. By measuring 5'UTR isoforms in retinal single-cell dataset, our work provides a comprehensive panorama of alternative TSS and its potential consequence related to post-transcriptional regulation. We anticipate our assay can not only provide insights into cellular heterogeneity driven by transcriptional initiation, but also open up the perspectives for identification of novel diagnostic indexes for diabetic retinopathy.

Osimertinib alone as second-line treatment for brain metastases (BM) control may be more limited than for non-BM in advanced NSCLC patients with an acquired EGFR T790M mutation.

BACKGROUND: This study was designed to investigate the difference between brain metastases (BM) and non-brain metastases (non-BM) treated by osimertinib in advanced patients with an acquired EGFR T790M mutation after obtaining first-generation EGFR-TKI resistance. METHODS: A total number of 135 first-generation EGFR-TKI-resistant patients with an acquired EGFR T790M mutation were retrospectively analyzed. The patients were divided into BM and non-BM groups. According to the type of treatment (whether brain radiotherapy), the BM patients were divided into an osimertinib combined with brain radiotherapy group and an osimertinib without brain radiotherapy group. In addition, according to the type of BM (the sequence between BM and osimertinib), the BM patients were subdivided into an osimertinib after BM group (initial BM developed after obtaining first-generation EGFR-TKI resistance) and an osimertinib before BM group (first-generation EGFR-TKI resistance then osimertinib administration performed; initial BM was not developed until osimertinib resistance). The progression-free survival (PFS) and overall survival (OS) were evaluated. The primary endpoint was OS between BM and no-BM patients. The secondary endpoints were PFS of osimertinib, and OS between brain radiotherapy and non-brain radiotherapy patients. RESULTS: A total of 135 patients were eligible and the median follow-up time of all patients was 50 months. The patients with BM (n = 54) had inferior OS than those without BM (n = 81) (45 months vs. 55 months, P = 0.004). And in BM group, the OS was longer in patients that received osimertinib combined with brain radiotherapy than in those without brain radiotherapy (53 months vs. 40 months, P = 0.014). In addition, the PFS was analysed according to whether developed BM after osimertinib resistance. The PFS of the patients that developed BM after acquiring osimertinib resistance was shorter than that without BM development, whether patients developed initial BM after first-generation EGFR-TKI resistance (7 months vs. 13 months, P = 0.003), or developed non-BM after first-generation EGFR-TKI resistance (13 months vs. 17 months, P < 0.001). CONCLUSIONS: In advanced patients with an acquired EGFR T790M mutation after obtaining first-generation EGFR-TKI resistance, osimertinib may be more limited in its control in BM than in non-BM. Also, osimertinib combined with brain radiotherapy may improve the survival time of BM patients.

Auxiliary diagnostic value of tumor biomarkers in pleural fluid for lung cancer-associated malignant pleural effusion.

BACKGROUND: Pleural effusion (PE) can be divided into benign pleural effusion (BPE) and malignant pleural effusion (MPE). There is no consensus on the identification of lung cancer-associated MPE using the optimal cut-off levels from five common tumor biomarkers (CEA, CYFRA 21-1, CA125, SCC-Ag, and NSE). Therefore, we aimed to find indicators for the auxiliary diagnosis of lung cancer-associated MPE by analyzing and then validating the optimal threshold levels of these biomarkers in pleural fluid (PF) and serum, as well as the PF/serum ratio. PATIENTS AND METHOD: The study has two sets of patients, i.e. the training set and the test set. In the training set, 348 patients with PE, between January 1, 2016 and December 31, 2017, were divided into BPE and MPE based on the cytological diagnosis. Subsequently, the optimal cut-off levels of tumor biomarkers were analyzed. In the test set, the diagnostic compliance rate was verified with 271 patients with PE from January 1, 2018 to July 31, 2019 to evaluate the auxiliary diagnostic value of the aforementioned indicators. RESULT: In the training set, PF CEA at the cut-off value of 5.23 ng/ml was the most effective indicator for MPE compared with other tumor biomarkers (all p < 0.001). In the test set, PF CEA at the cut-off value of 5.23 ng/ml showed the highest sensitivity, specificity and accuracy, positive and negative predictive value among other tumor biomarkers, which were 99.0%, 69.1%, 91.6%, 90.7%, and 95.9%, respectively. CONCLUSION: PF CEA at the cut-off level of 5.23 ng/ml was the most effective indicator for identifying lung cancer-associated MPE among the five common tumor biomarkers.

Integrated bioinformatics analysis of aberrantly-methylated differentially-expressed genes and pathways in age-related macular degeneration.

BACKGROUND: Age-related macular degeneration (AMD) represents the leading cause of visual impairment in the aging population. The goal of this study was to identify aberrantly-methylated, differentially-expressed genes (MDEGs) in AMD and explore the involved pathways via integrated bioinformatics analysis. METHODS: Data from expression profile GSE29801 and methylation profile GSE102952 were obtained from the Gene Expression Omnibus database. We analyzed differentially-methylated genes and differentially-expressed genes using R software. Functional enrichment and protein-protein interaction (PPI) network analysis were performed using the R package and Search Tool for the Retrieval of Interacting Genes online database. Hub genes were identified using Cytoscape. RESULTS: In total, 827 and 592 genes showed high and low expression, respectively, in GSE29801; 4117 hyper-methylated genes and 511 hypo-methylated genes were detected in GSE102952. Based on overlap, we categorized 153 genes as hyper-methylated, low-expression genes (Hyper-LGs) and 24 genes as hypo-methylated, high-expression genes (Hypo-HGs). Four Hyper-LGs (CKB, PPP3CA, TGFB2, SOCS2) overlapped with AMD risk genes in the Public Health Genomics and Precision Health Knowledge Base. KEGG pathway enrichment analysis indicated that Hypo-HGs were enriched in the calcium signaling pathway, whereas Hyper-LGs were enriched in sphingolipid metabolism. In GO analysis, Hypo-HGs were enriched in fibroblast migration, membrane raft, and coenzyme binding, among others. Hyper-LGs were enriched in mRNA transport, nuclear speck, and DNA binding, among others. In PPI network analysis, 23 nodes and two edges were established from Hypo-HGs, and 151 nodes and 73 edges were established from Hyper-LGs. Hub genes (DHX9, MAPT, PAX6) showed the greatest overlap. CONCLUSION: This study revealed potentially aberrantly MDEGs and pathways in AMD, which might improve the understanding of this disease.

Pigmented paravenous retinochoroidal atrophy: a case report.

BACKGROUND: Pigmented paravenous retinochoroidal atrophy (PPRCA) is an unusual retinal degeneration, and its performance on optical coherence tomography angiography (OCTA) is unclear. We report a Chinese female case of PPRCA and her OCTA features. CASE PRESENTATION: A 66-year-old female patient was referred to the author's center for gradual progressive loss of vision in both eyes and photophobia of 2 years duration. She reported having no family history of inherited ocular diseases. The funduscopic examination revealed bone-spicule pigmentation and retinochoroidal atrophy along the retinal veins. This patient was diagnosed with PPRCA which is a rare disease, uncommon in females, more commonly affecting the paravascular fundus. Noninvasive imaging techniques features of this patient was described, including ultra-wide field fundus autofluorescence, spectral domain optical coherence tomography (SD-OCT), OCTA (SSADA), etc. The en face OCTA images demonstrated areas of flow void beneath the retinal pigment epithelium-Bruch membrane layer suggestive of choriocapillaris hypoperfusion that corresponded with indocyanine green angiography (ICGA). Further studies should be conducted to clarify the relationship between choriocapillaris hypoperfusion and the development of PPRCA. CONCLUSIONS: The OCTA features in patients with PPRCA has not been described previously in the literature. This case might provide preliminary information regarding the pathophysiology of PPRCA and improve our understanding of the nature of this disease.

[Methodology comparison and influence factors analysis of epidermal growth factor receptor mutation detection].

OBJECTIVE: To compare two different methods in detecting epidermal growth factor receptor (EGFR) mutation status of non-small cell lung cancer (NSCLC) , and investing the influences of different conserving methods in DNA quantity, quality and detecting results of formalin-fixed paraffin embedded (FFPE) tissue samples. METHODS: One hundred and fifty FFPE samples of advanced NSCLC were collected in Cancer Institute/Hospital, Chinese Academy of Medical Sciences from November 2010 to August 2011. Both scorpions amplification refractory mutation system (scorpions ARMS) and direct sequencing were used to detect EGFR mutation in exon 18, 19, 20 and 21, together analyzed the differences between these two methods. Samples with inconsistent results were collected clinical treatment and survival information for further analysis. Extracted DNA from 30 FFPE samples and conserved at -20 °C for 3 years, meanwhile the same 30 FFPE samples DNA were extracted after conserving for 3 years. The DNA quantity, quality and testing results of the two conserving methods were compared. RESULTS: The detection success rate of Scorpions ARMS (100%) was higher than direct sequencing (87.3%), with statistical significance ( χ(2) = 20.28, P < 0.05). The consistent rate of the two methods was 84.7%, and there were no significant differences between these two methods, with a high consistence (Kappa = 0.738, P < 0.001) . The clinical treatment and survival status of 5 patients with different testing results were analysed, which were consistent with ARMS testing results. The quality of DNA extracted from FFPE tissues which were conserved for 3 years according to the conventional way was worse and with more fragments compared with DNA conserved at -20 °C for 3 years, but the EGFR mutation detecting results of the DNA acquired from the two conserved ways were consistent. CONCLUSIONS: Combination of Scorpions ARMS and direct sequencing could make the detecting result more comprehensive and reliable. Clinical treatment and survival data may provide diagnosis support for cases with different mutation testing results. DNA extracted from FFPE samples and stored at low temperature is better for conserving the integrity.

Intravitreal injection practice patterns among Chinese ophthalmologists.

AIM: To describe the practice patterns of intravitreal injections (IVIs) among ophthalmologists in China. METHODS: This was a cross-sectional online survey. Ophthalmologists who had performed accumulated more than 100 injections were contacted by the Brightness Center, a hospital-based national network, to complete an anonymous, 24-question, internet-based survey. They were surveyed on practices in injection techniques, pre-, and post-injections procedures. RESULTS: A total of 333 ophthalmologists from 28 provinces/municipalities/autonomous regions responded to the survey (50.68% response rate). The 91.29% of the respondents evaluated systemic risk factors by medical history, electrocardiogram (ECG) and blood test. All the respondents used pre-injection prophylactic antibiotics. Most checked intraocular pressure (IOP, 99.1%) and blood pressure (96.1%) before injections. A majority of the respondents performed injections in the operating room (98.8%), wore masks (99.7%), gloves (99.4%) and sterile surgical clothing (96.1%), performed topical anesthetics (97.9%), and applied povidone-iodine (95.8%) pre-injection. The 61.26% of the respondents dilated pupil. About half of the respondents (51.05%) performed bilateral injections in the same setting. Superior temporal quadrant (40.54%) was the most frequent site of injection. Around three quarters used 30-gauge needles. Most respondents (97.9%) measured the site of injection from limbus. More than half (53.45%) performed conjunctiva displacement prior to injection. The 32.43% of the respondents checked IOP post-injection and 87.99% physicians checked hand motion (HM) or counting fingers (CF) after injection, while 36.94% observed optic nerve perfusion. All participants used topical antibiotics post-injections. Most physicians (91.89%) reviewed patients on the following day. CONCLUSION: This study provides a description of the real-world practice patterns in IVIs in China and offers critical information regarding education and training of ophthalmologists and amendment of local society guidelines.

UCP2-SIRT3 Signaling Relieved Hyperglycemia-Induced Oxidative Stress and Senescence in Diabetic Retinopathy.

Purpose: Diabetic retinopathy (DR) is one of the most common reasons for blindness. uncoupling protein 2 (UCP2), an uncoupling protein located in mitochondria, has been reported to be related to metabolic and vascular diseases. This research aimed to illustrate the function and mechanism of UCP2 in the pathogenesis of DR. Methods: Human epiretinal membranes were collected to investigate the expression of UCP2 by quantitative real-time polymerase chain reaction (qRT-PCR) and immunofluorescence. Primary human retinal microvascular endothelial cells (HRECs) were cultured in high glucose (HG) to establish an in vitro cell model for DR. Flow cytometry analysis was used to measure intracellular reactive oxygen species (ROS). Senescence levels were evaluated by the senescence-associated beta-galactosidase (SA-ß-gal) assay, the expression of senescence marker P21, and cell-cycle analysis. Adenovirus-mediated UCP2 overexpression or knockdown and specific inhibitors were administered to investigate the underlying regulatory mechanism. Results: Proliferative fibrovascular membranes from patients with DR illustrated the downregulation of UCP2 and sirtuin 3 (SIRT3) by qRT-PCR and immunofluorescence. Persistent hyperglycemia-induced UCP2 downregulation in the progress of DR and adenovirus-mediated UCP2 overexpression protected endothelial cells from hyperglycemia-induced oxidative stress and senescence. Under hyperglycemic conditions, UCP2 overexpression attenuated NAD+ downregulation; hence, it promoted the expression and activity of SIRT3, an NAD+-dependent deacetylase regulating mitochondrial function. 3-TYP, a selective SIRT3 inhibitor, abolished the UCP2-mediated protective effect against oxidative stress and senescence. Conclusions: UCP2 overexpression relieved oxidative stress and senescence based on a novel mechanism whereby UCP2 can regulate the NAD+-SIRT3 axis. Targeting oxidative stress and senescence amelioration, UCP2-SIRT3 signaling may serve as a method for the prevention and treatment of DR and other diabetic vascular diseases.

Anti-angiogenic therapy or immunotherapy? A real-world study of patients with advanced non-small cell lung cancer with EGFR/HER2 exon 20 insertion mutations.

Background: For patients with EGFR/HER2 exon20 insertions, platinum-containing double-drug chemotherapy is still the standard treatment method. First-generation TKIs have almost no therapeutic activity against EGFR exon 20 insertions. The efficacy of second-and third-generation TKIs is still controversial. Immunotherapy research is scarce, and there is an urgent need for more evidence and new treatment options for this group of patients. Methods: We reviewed patients with advanced NSCLC with EGFR/HER2 exon 20 insertion mutations treated in Shanghai Chest Hospital and Shanghai Pulmonary Hospital from 2015 to 2022 and assessed the efficacy of receiving chemotherapy, anti-angiogenic therapy and immunotherapy, including objective response rate (ORR) and disease control rate (DCR), and compared progression-free survival (PFS) and overall survival (OS). Results: Of the 126 patients included in the study, 51 patients had EGFR20ins mutations and 7 5 patients had HER2-20ins mutations. In the first-line treatment, bevacizumab + chemotherapy (Beva+Chemo), ICI+chemotherapy (ICI+Chemo), compared with chemotherapy alone (Chemo), ORR: 40% vs 33.3% vs 15% (p=0.0168); DCR: 84% vs 80.9% vs 67.5% (p=0.1817); median PFS: 8.3 vs 7.0 vs 4.6 months (p=0.0032), ICI+Chemo has a trend of benefiting on OS. Stratified analysis showed that compared with chemotherapy, ICI+Chemo was more effective for EGFR20ins mutation with median PFS: 10.3 vs. 6.3m (P=0.013); Beva+Chemo was more effective for HER2-20ins mutation, with a median PFS: 6.6 vs. 4.3m (p=0.030). In the second-line treatment of EGFR20ins mutation, bevacizumab + chemotherapy has a significant advantage in PFS compared with targeted therapy, median PFS:10.8 vs 4.0 months (P=0.016). Conclusion: For patients with EGFR20ins mutation, compared to chemotherapy, ICI+Chemo prolongs PFS, and after chemotherapy progression, bevacizumab combined with chemotherapy seems better than Furmonertinib-based targeted therapy on PFS. For HER2-20ins mutation, Beva+Chemo may be a better choice.

Association of metabolomics with PD-1 inhibitor plus chemotherapy outcomes in patients with advanced non-small-cell lung cancer.

BACKGROUND: Combining immune checkpoint inhibitors (ICIs) with chemotherapy has become a standard treatment for patients with non-small cell lung cancer (NSCLC) lacking driver gene mutations. Reliable biomarkers are essential for predicting treatment outcomes. Emerging evidence from various cancers suggests that early assessment of serum metabolites could serve as valuable biomarkers for predicting outcomes. This study aims to identify metabolites linked to treatment outcomes in patients with advanced NSCLC undergoing first-line or second-line therapy with programmed cell death 1 (PD-1) inhibitors plus chemotherapy. METHOD: 200 patients with advanced NSCLC receiving either first-line or second-line PD-1 inhibitor plus chemotherapy, and 50 patients undergoing first-line chemotherapy were enrolled in this study. The 200 patients receiving combination therapy were divided into a Discovery set (n=50) and a Validation set (n=150). These sets were further categorized into respond and non-respond groups based on progression-free survival PFS criteria (PFS≥12 and PFS<12 months). Serum samples were collected from all patients before treatment initiation for untargeted metabolomics analysis, with the goal of identifying and validating biomarkers that can predict the efficacy of immunotherapy plus chemotherapy. Additionally, the validated metabolites were grouped into high and low categories based on their medians, and their relationship with PFS was analyzed using Cox regression models in patients receiving combination therapy. RESULTS: After the impact of chemotherapy was accounted for, two significant differential metabolites were identified in both the Discovery and Validation sets: N-(3-Indolylacetyl)-L-alanine and methomyl (VIP>1 and p<0.05). Notably, upregulation of both metabolites was observed in the group with a poorer prognosis. In the univariate analysis of PFS, lower levels of N-(3-Indolylacetyl)-L-alanine were associated with longer PFS (HR=0.59, 95% CI, 0.41 to 0.84, p=0.003), and a prolonged PFS was also indicated by lower levels of methomyl (HR=0.67, 95% CI, 0.47 to 0.96, p=0.029). In multivariate analyses of PFS, lower levels of N-(3-Indolylacetyl)-L-alanine were significantly associated with a longer PFS (HR=0.60, 95% CI, 0.37 to 0.98, p=0.041). CONCLUSION: Improved outcomes were associated with lower levels of N-(3-Indolylacetyl)-L-alanine in patients with stage IIIB-IV NSCLC lacking driver gene mutations, who underwent first-line or second-line therapy with PD-1 inhibitors combined with chemotherapy. Further exploration of the potential predictive value of pretreatment detection of N-(3-Indolylacetyl)-L-alanine in peripheral blood for the efficacy of combination therapy is warranted. STATEMENT: The combination of ICIs and chemotherapy has established itself as the new standard of care for first-line or second-line treatment in patients with advanced NSCLC lacking oncogenic driver alterations. Therefore, identifying biomarkers that can predict the efficacy and prognosis of immunotherapy plus chemotherapy is of paramount importance. Currently, the only validated predictive biomarker is programmed cell death ligand-1 (PD-L1), but its predictive value is not absolute. Our study suggests that the detection of N-(3-Indolylacetyl)-L-alanine in patient serum with untargeted metabolomics prior to combined therapy may predict the efficacy of treatment. Compared with detecting PD-L1 expression, the advantage of our biomarker is that it is more convenient, more dynamic, and seems to work synergistically with PD-L1 expression.

The prognosis and metabolite changes of NSCLC patients receiving first-line immunotherapy combined chemotherapy in different M1c categories according to 9th edition of TNM classification.

BACKGROUND: The 9th edition of the TNM Classification for lung cancer delineates M1c into two subcategories: M1c1 (Multiple extrathoracic lesions within a single organ system) and M1c2 (Multiple extrathoracic lesions involving multiple organ systems). Existing research indicates that patients with lung cancer in stage M1c1 exhibit superior overall survival compared to those in stage M1c2. The primary frontline therapy for patients with advanced non-small cell lung cancer (NSCLC), lacking driver gene mutations, involves the use of immune checkpoint inhibitors (ICIs) combined with chemotherapy. Nevertheless, a dearth of evidence exists regarding potential survival disparities between NSCLC patients with M1c1 and M1c2 undergoing first-line immune-chemotherapy, and reliable biomarkers for predicting treatment outcomes are elusive. Serum metabolic profiles may elucidate distinct prognostic mechanisms, necessitating the identification of divergent metabolites in M1c1 and M1c2 undergoing combination therapy. This study seeks to scrutinize survival discrepancies between various metastatic patterns (M1c1 and M1c2) and pinpoint metabolites associated with treatment outcomes in NSCLC patients undergoing first-line ICIs combined with chemotherapy. METHOD: In this study, 33 NSCLC patients lacking driver gene mutations diagnosed with M1c1, and 22 similarly diagnosed with M1c2 according to the 9th edition of TNM Classification, were enrolled. These patients received first-line PD-1 inhibitor plus chemotherapy. The relationship between metastatic patterns and progression-free survival (PFS) in patients undergoing combination therapy was analyzed using univariate and multivariate Cox regression models. Serum samples were obtained from all patients before treatment initiation for untargeted metabolomics analysis, aiming to identify differential metabolites. RESULTS: In the univariate analysis of PFS, NSCLC patients in M1c1 receiving first-line PD-1 inhibitor plus chemotherapy exhibited an extended PFS (HR = 0.49, 95% CI, 0.27-0.88, p = 0.017). In multivariate PFS analyses, these M1c1 patients receiving first-line PD-1 inhibitor plus chemotherapy also demonstrated prolonged PFS (HR = 0.45, 95% CI, 0.22-0.92, p = 0.028). The serum metabolic profiles of M1c1 and M1c2 undergoing first-line PD-1 inhibitors plus chemotherapy displayed notable distinctions. In comparison to M1c1 patients, M1c2 patients exhibited alterations in various pathways pretreatment, including platelet activation, linoleic acid metabolism, and the VEGF signaling pathway. Diminished levels of lipid-associated metabolites (diacylglycerol, sphingomyelin) were correlated with adverse outcomes. CONCLUSION: NSCLC patients in M1c1, devoid of driver gene mutations, receiving first-line PD-1 inhibitors combined with chemotherapy, experienced superior outcomes compared to M1c2 patients. Moreover, metabolomic profiles strongly correlated with the prognosis of these patients, and M1c2 patients with unfavorable outcomes manifested distinct changes in metabolic pathways before treatment. These changes predominantly involved alterations in lipid metabolism, such as decreased diacylglycerol and sphingomyelin, which may impact tumor migration and invasion.

Metadata information and fundus image fusion neural network for hyperuricemia classification in diabetes.

OBJECTIVE: In diabetes mellitus patients, hyperuricemia may lead to the development of diabetic complications, including macrovascular and microvascular dysfunction. However, the level of blood uric acid in diabetic patients is obtained by sampling peripheral blood from the patient, which is an invasive procedure and not conducive to routine monitoring. Therefore, we developed deep learning algorithm to detect noninvasively hyperuricemia from retina photographs and metadata of patients with diabetes and evaluated performance in multiethnic populations and different subgroups. MATERIALS AND METHODS: To achieve the task of non-invasive detection of hyperuricemia in diabetic patients, given that blood uric acid metabolism is directly related to estimated glomerular filtration rate(eGFR), we first performed a regression task for eGFR value before the classification task for hyperuricemia and reintroduced the eGFR regression values into the baseline information. We trained 3 deep learning models: (1) metadata model adjusted for sex, age, body mass index, duration of diabetes, HbA1c, systolic blood pressure, diastolic blood pressure; (2) image model based on fundus photographs; (3)hybrid model combining image and metadata model. Data from the Shanghai General Hospital Diabetes Management Center (ShDMC) were used to develop (6091 participants with diabetes) and internally validated (using 5-fold cross-validation) the models. External testing was performed on an independent dataset (UK Biobank dataset) consisting of 9327 participants with diabetes. RESULTS: For the regression task of eGFR, in ShDMC dataset, the coefficient of determination (R2) was 0.684±0.07 (95 % CI) for image model, 0.501±0.04 for metadata model, and 0.727±0.002 for hybrid model. In external UK Biobank dataset, a coefficient of determination (R2) was 0.647±0.06 for image model, 0.627±0.03 for metadata model, and 0.697±0.07 for hybrid model. Our method was demonstrably superior to previous methods. For the classification of hyperuricemia, in ShDMC validation, the area, under the curve (AUC) was 0.86±0.013for image model, 0.86±0.013 for metadata model, and 0.92±0.026 for hybrid model. Estimates with UK biobank were 0.82±0.017 for image model, 0.79±0.024 for metadata model, and 0.89±0.032 for hybrid model. CONCLUSION: There is a potential deep learning algorithm using fundus photographs as a noninvasively screening adjunct for hyperuricemia among individuals with diabetes. Meanwhile, combining patient's metadata enables higher screening accuracy. After applying the visualization tool, it found that the deep learning network for the identification of hyperuricemia mainly focuses on the fundus optic disc region.

Diagnostic Challenges in PD-1 Inhibitor Induced Panuveitis in a Teenage Girl with Chest Soft Tissue Sarcoma - A Case Report.

PURPOSE: To report a case of programmed cell death receptor-1 (PD-1) inhibitor induced panuveitis. METHOD: Observational case report of a 13-year-old Chinese girl presented as panuveitis. The clinical course, imaging performance, laboratory examination, differential diagnosis, treatment and prognosis were described. RESULT: Patient presented with bilateral anterior granulomatous uveitis, vitritis, papillitis, and various creamy yellow nodular lesions in the mid-peripheral fundus. She had a history of biopsy proven alveolar soft tissue sarcoma on the chest wall and pulmonary metastasis, and a PD-1 inhibitor (sintilimab) was intravenously administered. Blood tests, magnetic resonance imaging of the cranium and the orbit, aqueous humor assay of inflammatory cytokines and microbial DNA were performed to distinguish infectious and non-infectious uveitis, choroidal metastases, and intravenous injection-related endophthalmitis. The oncologist evaluated that the sarcoma was stable and terminated sintilimab dosage. After sintilimab withdrawal, the blurred vision improved. Then, the patient received oral corticosteroids, resulted in resolution of the panuveitis. A diagnosis of PD-1 inhibitor induced panuveitis was made. CONCLUSION: For patients taking PD-1 inhibitors, the major diagnostic challenge is to identify whether the cause of the uveitis is due to the antitumor treatment or not. It is suggested to be screened by eye care specialist and timely referral to uveitis specialist with any suspicion of intraocular inflammation for these patients.

Novel insights into the mechanisms of hard exudate in diabetic retinopathy: Findings of serum lipidomic and metabolomics profiling.

Objective: Retinal hard exudates (HEs) result from lipoproteins leaking from capillaries into extracellular retinal space, and are related to decreased visual acuity in diabetic retinopathy (DR). This study aims to identify differential serum lipids and metabolites associated with HEs. Materials and methods: A cross-sectional study was conducted Jul 2017 âˆ¼ Mar 2021. We assessed the amount of HEs using standard ETDRS photographs for comparison. HEs severity was rated as "no or questionable", "moderate" or "severe". Serum samples were processed via high coverage pseudotargeted lipidomics analysis, and untargeted liquid chromatography coupled with time-of-flight mass spectrometry for metabolomics study, respectively. Weighted gene co-expression network analyses, partial least squares-discriminant analysis, and multi-receiver operating characteristic analysis were applied. Results: A total of 167 patients were included. Discovery group: 116 eyes (116 patients). Validation group: 51 eyes (51 patients). 888 lipids were detected and divided into 18 modules (MEs), ME1 âˆ¼ ME18. Lipids in ME1 significantly increased in patients with HEs in DR (NPDR and PDR combined), NPDR, and PDR, respectively. ME1 enriched to triglycerides (29%), ceramides (17%), and N-acylethanolamines (15%). A combined model of 20 lipids was the best to discriminate HEs, area under curve = 0.804, 95% confidence interval = 0.674-0.916. For metabolomics analysis, 19 metabolites and 13 pathways associated with HEs were identified. Taurine and hypotaurine metabolism, cysteine and methionine metabolism were closely related to HEs (P < 0.01). Conclusions: The lipids and metabolites identified may serve as prediction biomarkers in the early stage of HEs in DR.

Single-cell RNA-sequencing uncovers the dynamic changes of tumour immune microenvironment in advanced lung adenocarcinoma.

BACKGROUND: The heterogeneity of lung adenocarcinoma (LUAD) plays a vital role in determining the development of cancer and therapeutic sensitivity and significantly hinders the clinical treatment of LUAD. OBJECTIVE: To elucidate the cellular composition and reveal previously uncharacterised tumour microenvironment in LUAD using single-cell RNA-sequencing (scRNA-seq). METHODS: Two scRNA-seq datasets with 106 829 high-quality cells from 34 patients including 11 normal, 9 early (stage I and II) and 14 advanced (stage III and IV) LUAD were integrated and clustered to explore diagnostic and therapeutic cell populations and their biomarkers for diverse stages of LUAD. Three independent bulk RNA-seq datasets were used to validate the results from scRNA-seq analysis. The expression of marker genes for specific cell types in early and advanced LUAD was verified by immunohistochemistry (IHC). RESULTS: Comprehensive cluster analysis identified that S100P+ epithelial and SPP1+ macrophage, positively related to poor outcomes, were preferentially enriched in advanced stage. Although the accumulation of KLRB1+CD8+ T cell and IGHA1+/IGHG1+ plasma cell both significantly associated the favourable prognosis, we also found KLRB1+CD8+ T cell decreased in advanced stage while IGHA1+/IGHG1+ plasma cells were increased. Cell-cell communication analysis showed that SPP1+ macrophage could interact with most of CD8+ subclusters through SPP1-CD44 axis. Furthermore, based on three independent bulk RNA-seq datasets, we built risk model with nine marker genes for specific cell subtypes and conducted deconvolution analysis, both supporting our results from scRNA-seq data. We finally validated the expression of four marker genes in early and advanced LUAD by IHC. CONCLUSION: Our analyses highlight the molecular dynamics of LUAD epithelial and microenvironment and provide new targets to improve LUAD therapy.

Association analysis between chromosomal abnormalities and fetal ultrasonographic soft markers based on 15,263 fetuses.

BACKGROUND: Soft markers are common prenatal ultrasonographic findings that indicate an increased risk for fetal aneuploidy. However, the association between soft markers and pathogenic or likely pathogenic copy number variations is still unclear, and clinicians lack clarity on which soft markers warrant a recommendation for invasive prenatal genetic testing of the fetus. OBJECTIVE: This study aimed to provide guidance on ordering prenatal genetic testing for fetuses with different soft markers and to elucidate the association between specific types of chromosomal abnormalities and specific ultrasonographic soft markers. STUDY DESIGN: Low-pass genome sequencing was performed for 15,263 fetuses, including 9123 with ultrasonographic soft markers and 6140 with normal ultrasonographic findings. The detection rate of pathogenic or likely pathogenic copy number variants among fetuses with various ultrasonographic soft markers were compared with that of fetuses with normal ultrasonography. The association of soft markers with aneuploidy and pathogenic or likely pathogenic copy number variants were investigated using Fisher exact tests with Bonferroni correction. RESULTS: The detection rate of aneuploidy and pathogenic or likely pathogenic copy number variants was 3.04% (277/9123) and 3.40% (310/9123), respectively, in fetuses with ultrasonographic soft markers. An absent or a hypoplastic nasal bone was the soft marker in the second trimester with the highest diagnostic rate for aneuploidy of 5.22% (83/1591) among all isolated groups. Four types of isolated ultrasonographic soft markers, namely a thickened nuchal fold, single umbilical artery, mild ventriculomegaly, and absent or hypoplastic nasal bone, had higher diagnostic rates for pathogenic or likely pathogenic copy number variants (P<.05; odds ratio, 1.69-3.31). Furthermore, this study found that the 22q11.2 deletion was associated with an aberrant right subclavian artery, whereas the 16p13.11 deletion, 10q26.13-q26.3 deletion, and 8p23.3-p23.1 deletion were associated with a thickened nuchal fold, and the 16p11.2 deletion and 17p11.2 deletion were associated with mild ventriculomegaly (P<.05). CONCLUSION: Ultrasonographic phenotype-based genetic testing should be considered in clinical consultations. Copy number variant analysis is recommended for fetuses with an isolated thickened nuchal fold, a single umbilical artery, mild ventriculomegaly, and an absent or a hypoplastic nasal bone. A comprehensive definition of genotype-phenotype correlations in aneuploidy and pathogenic or likely pathogenic copy number variants could provide better information for genetic counseling.

Effect and outcomes analysis of anlotinib in non-small cell lung cancer patients with liver metastasis: results from the ALTER 0303 phase 3 randomized clinical trial.

PURPOSE: Liver metastasis (LM) is common in non-small cell lung cancer (NSCLC), and always predicted worse outcomes with no effective therapy. We aimed to evaluate the effects and prognosis in LM patients treated with anlotinib. METHODS: The present study is a post hoc analysis based on a multicenter, double-blind, phase 3 randomized clinical trial which designed to evaluate the efficacy and safety of anlotinib in patients with advanced NSCLC. A total of 437 patients were enrolled in present study, and 78 patients with LM. RESULTS: Patients with LM showed a worse outcome compared to those without LM (PFS median, 2.6 vs 4.2 months), and OS (median, 5.6 vs 9.4 months, both P < 0.0001). The anlotinib was associated with longer PFS (median, 3.0 months) compared with placebo (median, 0.9 months), with a hazard ratio (HR) of 0.23 (95%CI, 0.12-0.42; P < 0.0001). Furthermore, OS was marginally significantly better in anlotinib group (median 6.6 months), compared with placebo (median 4.0 months), HR 0.61 (95%CI, 0.36-1.02; P = 0.055). Multivariate analysis confirmed normal peripheral blood LDH/TBiL level predicted better PFS and OS, lower ECOG score acted as independently prognostic factor for superior OS. Anlotinib was more associated with hand-foot syndrome (7.7% vs 0) and serum TSH level rise (7.7% vs 3.8%) and well tolerated, all AEs were no more than grade 3. CONCLUSION: Patients with LM had a dismal prognosis, anlotinib could lead to a better PFS in pretreated NSCLC patients, which suggested anlotinib is a potential third-line or further therapy in these patients.