The Acetic Acid Produced by Lactobacillus Species Regulates Immune Function to Alleviate PEDV Infection in Piglets.

Porcine epidemic diarrhea virus (PEDV) infection results in significant mortality among newborn piglets, leading to substantial economic setbacks in the pig industry. Short-chain fatty acids (SCFA), the metabolites of intestinal probiotics, play pivotal roles in modulating intestinal function, enhancing the intestinal barrier, and bolstering immune responses through diverse mechanisms. The protective potential of Lactobacillus delbrueckii, Lactobacillus johnsonii, and Lactococcus lactis was first noted when administered to PEDV-infected piglets. Histological evaluations, combined with immunofluorescence studies, indicated that piglets receiving L. lactis displayed less intestinal damage, with diminished epithelial cell necrosis and milder injury levels. Differences in immunofluorescence intensity revealed a significant disparity in antigen content between the L. lactis and PEDV groups, suggesting that L. lactis might suppress PEDV replication, the intestine. We then assessed short-chain fatty acid content through targeted metabolomics, finding that acetate levels markedly varied from other groups. This protective impact was confirmed by administering acetate to PEDV-infected piglets. Data suggested that piglets receiving acetate exhibited resistance to PEDV. Flow cytometry analyses were conducted to evaluate the expression of innate and adaptive immune cells in piglets. Sodium acetate appeared to bolster innate immune defenses against PEDV, marked by elevated NK cell and macrophage counts in mesenteric lymph nodes, along with increased NK cells in the spleen and macrophages in the bloodstream. Acetic acid was also found to enhance the populations of CD8+ IFN-γ T cells in the blood, spleen, and mesenteric lymph, CD4+ IFN-γ T cells in mesenteric lymph nodes and spleen, and CD4+ IL-4+T cells in the bloodstream. Transcriptome analyses were carried out on the jejunal mucosa from piglets with PEDV-induced intestinal damage and from healthy counterparts with intact barriers. Through bioinformatics analysis, we pinpointed 189 significantly upregulated genes and 333 downregulated ones, with the PI3K-AKT, ECM-receptor interaction, and pancreatic secretion pathways being notably enriched. This transcriptomic evidence was further corroborated by western blot and qPCR. Short-chain fatty acids (SCFA) were found to modulate G protein-coupled receptor 41 (GPR41) and 43 (GPR43) in porcine intestinal epithelial cells (IPEC-J2). Post-acetic acid exposure, there was a notable upsurge in the ZO-1 barrier protein expression in IPEC-J2 compared to the unexposed control group (WT), while GPR43 knockdown inversely affected ZO-1 expression. Acetic acid amplified the concentrations of phosphorylated PI3K and AKT pivotal components of the PI3K/AKT pathway. Concurrently, the co-administration of AKT agonist SC79 and PI3K inhibitor LY294002 revealed acetic acid's role in augmenting ZO-1 expression via the P13K/AKT signaling pathway. This study demonstrates that acetic acid produced by Lactobacillus strains regulates intestinal barrier and immune functions to alleviate PEDV infection. These findings provide valuable insights for mitigating the impact of PEDV in the pig industry.

Porous Se@SiO2 nanospheres alleviate diabetic retinopathy by inhibiting excess lipid peroxidation and inflammation.

BACKGROUND: Lipid peroxidation is a characteristic metabolic manifestation of diabetic retinopathy (DR) that causes inflammation, eventually leading to severe retinal vascular abnormalities. Selenium (Se) can directly or indirectly scavenge intracellular free radicals. Due to the narrow distinction between Se's effective and toxic doses, porous Se@SiO2 nanospheres have been developed to control the release of Se. They exert strong antioxidant and anti-inflammatory effects. METHODS: The effect of anti-lipid peroxidation and anti-inflammatory effects of porous Se@SiO2 nanospheres on diabetic mice were assessed by detecting the level of Malondialdehyde (MDA), glutathione peroxidase 4 (GPX4), decreased reduced/oxidized glutathione (GSH/GSSG) ratio, tumor necrosis factor (TNF)-α, interferon (IFN)-γ, and interleukin (IL) -1ß of the retina. To further examine the protective effect of porous Se@SiO2 nanospheres on the retinal vasculopathy of diabetic mice, retinal acellular capillary, the expression of tight junction proteins, and blood-retinal barrier destruction was observed. Finally, we validated the GPX4 as the target of porous Se@SiO2 nanospheres via decreased expression of GPX4 and detected the level of MDA, GSH/GSSG, TNF-α, IFN-γ, IL -1ß, wound healing assay, and tube formation in high glucose (HG) cultured Human retinal microvascular endothelial cells (HRMECs). RESULTS: The porous Se@SiO2 nanospheres reduced the level of MDA, TNF-α, IFN-γ, and IL -1ß, while increasing the level of GPX4 and GSH/GSSG in diabetic mice. Therefore, porous Se@SiO2 nanospheres reduced the number of retinal acellular capillaries, depletion of tight junction proteins, and vascular leakage in diabetic mice. Further, we identified GPX4 as the target of porous Se@SiO2 nanospheres as GPX4 inhibition reduced the repression effect of anti-lipid peroxidation, anti-inflammatory, and protective effects of endothelial cell dysfunction of porous Se@SiO2 nanospheres in HG-cultured HRMECs. CONCLUSION: Porous Se@SiO2 nanospheres effectively attenuated retinal vasculopathy in diabetic mice via inhibiting excess lipid peroxidation and inflammation by target GPX4, suggesting their potential as therapeutic agents for DR.

Gut microbiota-derived LCA mediates the protective effect of PEDV infection in piglets.

BACKGROUND: The gut microbiota is a critical factor in the regulation of host health, but the relationship between the differential resistance of hosts to pathogens and the interaction of gut microbes is not yet clear. Herein, we investigated the potential correlation between the gut microbiota of piglets and their disease resistance using single-cell transcriptomics, 16S amplicon sequencing, metagenomics, and untargeted metabolomics. RESULTS: Porcine epidemic diarrhea virus (PEDV) infection leads to significant changes in the gut microbiota of piglets. Notably, Landrace pigs lose their resistance quickly after being infected with PEDV, but transplanting the fecal microbiota of Min pigs to Landrace pigs alleviated the infection status. Macrogenomic and animal protection models identified Lactobacillus reuteri and Lactobacillus amylovorus in the gut microbiota as playing an anti-infective role. Moreover, metabolomic screening of the secondary bile acids' deoxycholic acid (DCA) and lithocholic acid (LCA) correlated significantly with Lactobacillus reuteri and Lactobacillus amylovorus, but only LCA exerted a protective function in the animal model. In addition, LCA supplementation altered the distribution of intestinal T-cell populations and resulted in significantly enriched CD8+ CTLs, and in vivo and in vitro experiments showed that LCA increased SLA-I expression in porcine intestinal epithelial cells via FXR receptors, thereby recruiting CD8+ CTLs to exert antiviral effects. CONCLUSIONS: Overall, our findings indicate that the diversity of gut microbiota influences the development of the disease, and manipulating Lactobacillus reuteri and Lactobacillus amylovorus, as well as LCA, represents a promising strategy to improve PEDV infection in piglets. Video Abstract.

Small-molecule agonist AdipoRon alleviates diabetic retinopathy through the AdipoR1/AMPK/EGR4 pathway.

BACKGROUND: Diabetes mellitus (DM) is a progressive disease that involves multiple organs due to increased blood glucose, and diabetic retinopathy (DR) is the main complication of DM in the eyes and causes irreversible vision loss. In the pathogenesis of diabetic vascular disease, oxidative stress caused by hyperglycemia plays an important role in Müller cell impairment. In recent years, AdipoRon, an adiponectin analog that demonstrated important physiological functions in obesity, diabetes, inflammation, and cardiovascular diseases, demonstrated cellular protection from apoptosis and reduced inflammatory damage through a receptor-dependent mechanism. Here, we investigated how AdipoRon reduced oxidative stress and apoptosis in Müller glia in a high glucose environment. RESULTS: By binding to adiponectin receptor 1 on Müller glia, AdipoRon activated 5' adenosine monophosphate-activated protein kinase (AMPK)/acetyl-CoA carboxylase phosphorylation downstream, thereby alleviating oxidative stress and eventual apoptosis of cells and tissues. Transcriptome sequencing revealed that AdipoRon promoted the synthesis and expression of early growth response factor 4 (EGR4) and inhibited the cellular protective effects of AdipoRon in a high-glucose environment by reducing the expression of EGR4. This indicated that AdipoRon played a protective role through the EGR4 and classical AMPK pathways. CONCLUSIONS: This provides a new target for the early treatment of DR.

Ginkgolide attenuates memory impairment and neuroinflammation by suppressing the NLRP3/caspase-1 pathway in Alzheimer's disease.

The NLRP3 inflammasome is involved in the neuroinflammatory pathway of Alzheimer's disease (AD). The aim of this study is to explore the roles and underlying mechanisms of ginkgolide (Baiyu®) on amyloid precursor protein (APP)/presenilin 1 (PS1) transgenic mice and a murine microglial cell line, BV-2. In the present study, the APP/PS1 mice were administered with ginkgolide, followed by a Morris water maze test. The mice were then euthanized to obtain brain tissue for histological and Aß analysis. Additionally, BV-2 cells were pretreated with ginkgolide and then incubated with Aß1-42 peptide. NLRP3, ASC, and caspase-1 mRNA and protein expression in brain tissue of mice and BV-2 cells were quantified by real-time PCR and western blotting, as well as reactive oxygen species (ROS) production, interleukin (IL)-1ß and IL-18 levels by lucigenin technique and ELISA. Compared with the APP/PS1 mice, ginkgolide-treated mice demonstrated the shortened escape latency, reduced plaques, less inflammatory cell infiltration and neuron loss in the hippocampi of APP/PS1 mice. The levels of NLRP3, ASC, caspase-1, ROS, IL-1ß, and IL-18 were also decreased in the brain tissue of APP/PS1 mice or Aß1-42-treated BV-2 cells following ginkgolide treatment. Ginkgolide exerted protective effects on AD, at least partly by inactivating the NLRP3/caspase-1 pathway.

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.

Serum Ang-1/Ang-2 ratio may be a promising biomarker for evaluating severity of diabetic retinopathy.

PURPOSE: To investigate the predictive role of serum angiopoietin-1 and angiopoietin-2 (Ang-1/Ang-2) in evaluating the severity of diabetic retinopathy (DR). METHODS: A total of 101 outpatients with type 2 diabetes mellitus (T2DM) were recruited and were further divided into the following five groups: T2DM without DR (non-DR), mild non-proliferative DR (NPDR), moderate NPDR, severe NPDR and proliferative DR (PDR) in accordance with the International Clinical Diabetic Retinopathy Guidelines. Furthermore, 101 serum samples were included in the further analysis using enzyme-linked immunosorbent assays. A receiver operating characteristic (ROC) curve was plotted to evaluate the diagnostic value of each index. RESULTS: The expression of Ang-1 in the PDR group was significantly lower than that in the non-DR group, while Ang-2 showed an opposite upward trend (p < 0.05). The Ang-1/Ang-2 ratio of the non-DR group was significantly lower than that of the moderate NPDR, severe NPDR and PDR (p < 0.05, p < 0.01 and p < 0.01, respectively). Differences in the Ang-1/Ang-2 ratio were observed earlier than those in the individual Ang-1 and Ang-2 measurements. The maximal Youden index was 0.512 with a calculated area under the curve (AUC) value of 0.734 (p < 0.01). CONCLUSIONS: The Ang-1/Ang-2 ratio was helpful in assessing the severity of DR and may provide potential clinical benefits as a biomarker and early warning signs for DR diagnosis.

Clinical features, risk factors and survival in cardiac myxoma-related ischemic stroke: A multicenter case-control study.

BACKGROUND: Cardiac myxoma (CM) is an important etiology of stroke in young adults, but studies on CM-related ischemic stroke (CM-IS) are limited and conflicting. Hence, we investigated clinical characterizations, risk factors of CM-IS, and short-term survival after surgical resection. METHODS: We performed a retrospective analysis of data from all CM patients at three referral management centers and conducted follow-up examination. RESULTS: Among 414 CM patients, 402 were recruited for further analysis, including 54 patients with CM-IS and 348 patients with CM without stroke (Non-stroke). In the acute phase, patients presented with NIHSS 3 (interquartile range: 0-10) and clinical presentation comprising neurological, cardiac and constitutional symptoms. Multivariate analysis showed that the factors associated with an increased risk of CM-IS were tumor width < 30 mm [OR = 2.652, 95% CI: 1.061-6.627, P = 0.037], tumors with high-mobility (OR = 2.700, 95% CI: 1.357-5.371, P = 0.005), thrombus on the tumor surface (OR = 1.856, 95% CI: 1.003-3.434, P = 0.049), and lower B-type natriuretic peptide (BNP) levels (OR = 0.995, 95% CI: 0.989-0.999, P = 0.047). The overall three-year survival rate was 95.7% (95% CI: 94.9-96.5) in CM-IS patients who underwent surgery. CONCLUSIONS: CM-IS patients had mild or moderate neurologic deficits with various presentations at disease onset. Narrower tumor width, tumors with high-mobility, thrombus on the tumor surface, and lower BNP levels are potential predictors of CM-IS development. Surgical removal of CM is safe and efficacious in patients with CM-IS.

Characterization and pathogenicity of the porcine epidemic diarrhea virus isolated in China.

Piglet diarrhea caused by the porcine epidemic diarrhea virus (PEDV) is a common problem on pig farms in China associated with high morbidity and mortality rates. In this study, three PEDV isolates were successfully detected after the fourth blind passage in Vero cells. The samples were obtained from infected piglet farms in Jilin (Changchun), and Shandong (Qingdao) Provinces of China and were designated as CH/CC-1/2018, CH/CC-2/2018, and CH/QD/2018. According to the analysis of the complete S protein gene sequence, the CH/CC-1/2018 and CH/CC-2/2018 were allocated to the G2b branch, while CH/QD/2018 was located in the G1a interval and was closer to the vaccine strain CV777. Successful detection and identification of the isolated strains were carried out using electron microscopy and indirect immunofluorescence. Meanwhile, animal challenge experiments and viral RNA copies determination were used to compare the pathogenicity. The results showed that CH/CC-1/2018 in Changchun was more pathogenic than CH/QD/2018 in Qingdao. In conclusion, the discovery of these new strains is conducive to the development of vaccines to prevent the pandemic of PEDV, especially that the CH/CC-1/2018, and CH/CC-2/2018 were not related to the classical vaccine strain CV777.

Risk factors and a Bayesian network model to predict ischemic stroke in patients with dilated cardiomyopathy.

Objective: This study aimed to identify risk factors and create a predictive model for ischemic stroke (IS) in patients with dilated cardiomyopathy (DCM) using the Bayesian network (BN) approach. Materials and methods: We collected clinical data of 634 patients with DCM treated at three referral management centers in Beijing between 2016 and 2021, including 127 with and 507 without IS. The patients were randomly divided into training (441 cases) and test (193 cases) sets at a ratio of 7:3. A BN model was established using the Tabu search algorithm with the training set data and verified with the test set data. The BN and logistic regression models were compared using the area under the receiver operating characteristic curve (AUC). Results: Multivariate logistic regression analysis showed that hypertension, hyperlipidemia, atrial fibrillation/flutter, estimated glomerular filtration rate (eGFR), and intracardiac thrombosis were associated with IS. The BN model found that hyperlipidemia, atrial fibrillation (AF) or atrial flutter, eGFR, and intracardiac thrombosis were closely associated with IS. Compared to the logistic regression model, the BN model for IS performed better or equally well in the training and test sets, with respective accuracies of 83.7 and 85.5%, AUC of 0.763 [95% confidence interval (CI), 0.708-0.818] and 0.822 (95% CI, 0.748-0.896), sensitivities of 20.2 and 44.2%, and specificities of 98.3 and 97.3%. Conclusion: Hypertension, hyperlipidemia, AF or atrial flutter, low eGFR, and intracardiac thrombosis were good predictors of IS in patients with DCM. The BN model was superior to the traditional logistic regression model in predicting IS in patients with DCM and is, therefore, more suitable for early IS detection and diagnosis, and could help prevent the occurrence and recurrence of IS in this patient cohort.

Lactiplantibacillus plantarum 0111 Protects Against Influenza Virus by Modulating Intestinal Microbial-Mediated Immune Responses.

There are some limitations of traditional influenza vaccines concerning novel mutant strains. Therefore, it is particularly important to develop preventive means for antigen-unrelated types of influenza viruses. Recent studies have shown that probiotics can modulate the immune system and reduce the severity of viral infections. In this study, we investigated the potential of Lactiplantibacillus plantarum 0111 against influenza virus H9N2. Challenge experiments showed that L. plantarum 0111 pretreatments could effectively improve mice's survival rate and weight loss and reduce the inflammatory cytokines IL-6 and TNF-α in the lungs and bronchoalveolar lavage fluid (BALF) along with the degree of lung and intestinal injury. FMT experiment demonstrates that the protective effect produced by L. plantarum 0111 is associated with gut microorganisms. In addition, 16S high-throughput sequencing of the mouse intestinal microbiota showed that L. plantarum 0111 remodeled the intestinal microbiota after H9N2 infection and maintained the gut microbiota balance. In a mouse model, the oral administration of L. plantarum 0111 increased IFN-ß expression in the serum and BALF. At the same time, the transcript levels of IFN-ß and related ISGs in the intestine and lungs of mice were also increased. In addition, the activation and polarization of T cells in mesenteric lymph nodes (MLNs) and the spleen were detected by flow cytometry, and the results showed that L. plantarum 0111 modulated cytokines in T cells and increased IgA expression in B cells in the MLNs and spleen. Thus, L. plantarum 0111 may improve gut microbiota-mediated immune responses and thus, resist infection by the influenza virus, and it could be used as an effective preventive measure against the influenza virus.

Regulation of progesterone during follicular development by FSH and LH in sheep.

Progesterone (P4) can participate in the development of female mammalian antral follicles through nuclear receptor (PGR). In this experiment, the differences of P4 synthesis and PGR expression in different developmental stages of sheep antral follicles (large > 5mm, medium 2-5mm, small < 2mm) were detected by enzyme-linked immunosorbent assay, immunohistochemistry, qRT-PCR and Western blotting. Secondly, sheep follicular granulosa cells were cultured in vitro. The effects of different concentrations of FSH and LH on P4 synthesis and PGR expression were studied. The results showed that acute steroid regulatory protein (StAR), cholesterol side chain lyase (P450scc) and 3ß Hydroxysteroid dehydrogenase (3ß-HSD) and PGR were expressed in antral follicles, and with the development of antral follicles in sheep, StAR, P450scc and the expression of 3ß-HSD and PGR increased significantly. In vitro experiments showed that FSH and LH alone or together treatment could regulate P4 secretion and PGR expression in sheep follicular granulosa cells to varying degrees, hint P4 and PGR by FSH and LH, and LH was the main factor. Our results supplement the effects of FSH and LH on the regulation of P4 synthesis during follicular development, which provides new data for further study of steroid synthesis and function in follicular development.

Protective effect of pentraxin 3 on pathological retinal angiogenesis in an in vitro model of diabetic retinopathy.

BACKGROUND: Diabetic retinopathy (DR) is the most common retinal microvascular disease caused by diabetes. Previous studies indicated that Pentraxin 3 (PTX3), an acute phase reactant, was closely related to the development of DR. But the exact effect of PTX3 in diabetic retinopathy needs more investigations. METHODS: Real-time fluorescence quantitative polymerase chain reaction (qRT-PCR) analysis and western blot (WB) were used to detect the expression of PTX3 in vitro. The Ki67 immunofluorescent staining, scratch-wound migration assay, and tube formation experiments were performed to detect the effect of PTX3 knockdown and overexpression on the fibroblast growth factor (FGF)-induced proliferation, migration and tube-forming ability of human retinal microvascular endothelial cells (HRMECs). The phosphorylation levels of extracellular regulated protein kinases (ERK) and fibroblast growth factor receptor (FGFR) in HRMECs were detected by WB. RESULTS: In vitro, the mRNA and protein expressions of PTX3 in the high-concentration glucose condition group were upregulated compared with the normal group (p < 0.05). The proliferation, migration and tube-forming abilities of HRMECs exposed to high-concentration glucose were enhanced (p < 0.01, p < 0.01, p < 0.05 respectively), and the phosphorylation of FGFR and ERK1/2 were increased (p < 0.01, p < 0.05 respectively) compared with the normal condition group. Compared with the high glucose condition group, the proliferation, migration and tube-forming abilities of HRMECs in the high glucose + PTX3 siRNA condition group were further strengthened (p < 0.001, p < 0.0001, p < 0.05 respectively), and the phosphorylation of FGFR and ERK1/2 were increased (p < 0.001, p < 0.01 respectively). Compared with the high glucose condition group, the proliferation, migration and tube-forming abilities of HRMECs in the high glucose + PTX3 overexpression condition group were compromised (p < 0.001, p < 0.05, p < 0.01 respectively), and the phosphorylation of FGFR and ERK1/2 were inhibited (p < 0.001, p < 0.0001 respectively). Neither the scramble siRNA condition group nor the blank plasmid condition group showed significant difference on the proliferation, migration and tube-forming abilities of HRMECs compared with the high glucose condition group (p > 0.05). CONCLUSIONS: The upregulated expression of PTX3 may play a protective role on pathological angiogenesis in DR. PTX3 may serve as a new target for the treatment of DR.

Preliminary study on the differentiation of vulnerable carotid plaques via analysis of calcium content and spectral curve slope by using gemstone spectral imaging.

Growing evidence indicates that vulnerable carotid plaque rupture is an important cause of stroke. However, the role of novel gemstone spectral imaging (GSI) in the assessment of vulnerable carotid plaques has remained to be sufficiently explored. Therefore, the aim of the present study was to provide a comprehensive evaluation of carotid atherosclerotic plaques using both GSI imaging biomarkers and serological biomarkers, and further explore their possible roles in the atherogenic process. The present study analyzed GSI data, including calcium content of carotid atherosclerotic plaques and spectral curve slope, as well as serum high-sensitivity C-reactive protein (Hs-CRP) and monocyte chemotactic protein-1 (MCP-1) levels in patients with a carotid atherosclerotic plaque using GSI-computed tomographic angiography and immunoturbidimetry. Patients with unstable plaque exhibited a significantly lower calcium content and higher spectral curve slope than those of the stable plaque group. In addition, patients with unstable plaque exhibited an increase in Hs-CRP and MCP-1 levels compared with those of the stable plaque and normal control groups. The alteration in GSI calcium content and spectral curve slope reflects a close link between calcification and plaque instability, while aberrant Hs-CRP and MCP-1 expression are involved in the formation or development of vulnerable plaques. Taken together, the present results strongly support the feasibility of using these serological and newly identified imaging parameters as multiple potential biomarkers relevant to plaque vulnerability or stroke progression.

Protective Effects of Ginkgolide on a Cellular Model of Alzheimer's Disease via Suppression of the NF-κB Signaling Pathway.

SYNOPSIS: NF-κB signaling has been reported to play a key regulatory role in the pathogenesis of Alzheimer's disease (AD). The purpose of this study is to investigate the effects of ginkgolide on cell viability in an AD cellular model involving an APP/PS1 double gene-transfected HEK293 cell line (APP/PS1-HEK293) and further explore the mechanisms of action related to NF-κB signaling. The optimal time point and concentration of ginkgolide for cell proliferation were screened using a cell counting kit-8 assay. Based on the results, an in vitro study was performed by co-culture of APP/PS1-HEK293 with different dosages of ginkgolide, followed by an enzyme-linked immunosorbent assay to measure the levels of supernatant tumor necrosis factor (TNF)-α, interleukin (IL)-1ß and IL-6, as well as western blotting and real-time polymerase chain reaction to detect intracellular protein and mRNA expression of NF-κB p65, IκBa, Bcl-2, and Bax. APP/PS1-HEK293 cells exhibited the highest cell viability at a concentration of 100 µg/ml after 48 h of treatment with ginkgolide. The supernatant levels of TNF-α, IL-1ß, and IL-6 in the high-dosage ginkgolide-treated groups were lower than those in the control group. Compared with the control group, there were decreased intracellular protein and mRNA expression of NF-κB p65 and Bax, but increased protein and mRNA expression of IκBa in both high-dosage and low-dosage groups. Ginkgolide may enhance cell viability, indicative of its neuroprotective effects on AD, at least partially via suppression of the NF-κB signaling pathway involving anti-apoptosis and anti-inflammation mechanisms. Therefore, ginkgolide might be a promising therapeutic agent against AD.

A CRISPR-Based Method for Constructing Conditional Mutations of Essential Genes in Cyanobacteria.

Cyanobacteria, a group of diverse bacteria capable of oxygenic photosynthesis, are excellent models for investigating many important cellular processes, such as photosynthesis, nitrogen fixation, and prokaryotic cell differentiation. They also have great potential to become the next-generation cell factories for sustainable biosynthesis of valuable products. However, genetic manipulation in cyanobacteria is not as convenient as in other model bacteria. Particularly, handling essential genes in cyanobacteria has been difficult due to the lack of appropriate tools, limiting our understanding of many important cellular functions encoded by them. We recently develop a CRISPR-based method for constructing the conditional mutants of cyanobacterial essential genes by engineering the ribosome binding site to a theophylline-responsive riboswitch. Here, we provide the details of this method. The principle of this method could be used to construct conditional mutants in a wide range of bacterial species.

Noncovalent CDK12/13 dual inhibitors-based PROTACs degrade CDK12-Cyclin K complex and induce synthetic lethality with PARP inhibitor.

Cyclin-dependent kinase 12 (CDK12) plays a crucial role in DNA-damage response gene transcription and has recently been validated as a promising target in cancer therapy. However, existing CDK12 inhibitors potently inhibit its closest isoform CDK13, which could cause potential toxicity. Therefore, the development of CDK12 inhibitors with isoform-selectivity against CDK13 continues to be a challenge. By taking advantage of the emerging PROteolysis-TArgeting Chimeras (PROTACs) approach, we have synthesized a potent PROTAC degrader PP-C8 based on the noncovalent dual inhibitors of CDK12/13 and demonstrated its specificity for CDK12 over CDK13. Notably, PP-C8 induces profound degradation of cyclin K simultaneously and downregulates the mRNA level of DNA-damage response genes. Global proteomics profiling revealed PP-C8 is highly selective toward CDK12-cyclin K complex. Importantly, PP-C8 demonstrates profound synergistic antiproliferative effects with PARP inhibitor in triple-negative breast cancer (TNBC). The potent and selective CDK12 PROTAC degrader developed in this study could potentially be used to treat CDK12-dependent cancers as combination therapy.

A preliminary study of association of cigarette smoking with risk of neuromyelitis optica spectrum disorder.

ABSTRACT: Various studies have revealed an association between cigarette smoking and increased risk for multiple sclerosis (MS). However, its role in neuromyelitis optica spectrum disorder (NMOSD) remains elusive. Therefore, in the present case-control study, we aimed to assess the association of active and passive cigarette smoking with the risk of MS and NMOSD.Thirty-six patients with NMOSD, 46 patients with MS, and 122 healthy individuals were included in this study. Standardized questionnaires and telephone interviews were used to collect information regarding the active and passive cigarette smoking behaviors of the patients and normal controls.The risk of MS was significantly higher among smokers than among nonsmokers (odds ratio = 2.166, 95% confidence interval: 1.109-4.170; P = .027). Further analysis of the risk between active and passive smokers, male smokers and nonsmokers showed no statistical difference. However, neither smokers nor active smokers had a greater or lower risk of NMOSD than their nonsmoking counterparts.Our preliminary study showed no significant association between cigarette smoking and the risk of NMOSD, strongly suggesting that, unlike MS, cigarette smoking might not confer NMOSD susceptibility, at least in the Northern Han Chinese population.

Lamiophlomis herba: A comprehensive overview of its chemical constituents, pharmacology, clinical applications, and quality control.

Lamiophlomis rotata (Benth.) Kudo (LR) is an extensively used Chinese herbal medicine. It contains a variety of chemical constituents with significant biological activities that were first recorded in the classical masterpiece of Tibetan Medicine, Somaratsa. In this review, we summarize the information regarding the traditional uses, chemical constituents, pharmacological effects, clinical applications, quality control, toxicology, and pharmacokinetics of LR. At least 223 chemical constituents have been isolated from LR, including phenylethanoid glycosides, flavonoids, iridoids, volatile oils, et al. Their various physiological activities have been demonstrated as analgesic, hemostatic, anti-inflammatory, anti-tumor, marrow-supplementing, anti-bacterial, and immunity-strengthening. The clinical applications of LR and quality control are also discussed, as well as some existing problems. This article aims to provide more comprehensive information on the chemical composition, pharmacological activity, and clinical application of LR, so as to provide a theoretical basis for the further reasonable development of LR in clinical practice and of new drugs.