Isaridin E Protects against Sepsis by Inhibiting Von Willebrand Factor-Induced Endothelial Hyperpermeability and Platelet-Endothelium Interaction.

Endothelial hyperpermeability is pivotal in sepsis-associated multi-organ dysfunction. Increased von Willebrand factor (vWF) plasma levels, stemming from activated platelets and endothelium injury during sepsis, can bind to integrin αvß3, exacerbating endothelial permeability. Hence, targeting this pathway presents a potential therapeutic avenue for sepsis. Recently, we identified isaridin E (ISE), a marine-derived fungal cyclohexadepsipeptide, as a promising antiplatelet and antithrombotic agent with a low bleeding risk. ISE's influence on septic mortality and sepsis-induced lung injury in a mouse model of sepsis, induced by caecal ligation and puncture, is investigated in this study. ISE dose-dependently improved survival rates, mitigating lung injury, thrombocytopenia, pulmonary endothelial permeability, and vascular inflammation in the mouse model. ISE markedly curtailed vWF release from activated platelets in septic mice by suppressing vesicle-associated membrane protein 8 and soluble N-ethylmaleide-sensitive factor attachment protein 23 overexpression. Moreover, ISE inhibited healthy human platelet adhesion to cultured lipopolysaccharide (LPS)-stimulated human umbilical vein endothelial cells (HUVECs), thereby significantly decreasing vWF secretion and endothelial hyperpermeability. Using cilengitide, a selective integrin αvß3 inhibitor, it was found that ISE can improve endothelial hyperpermeability by inhibiting vWF binding to αvß3. Activation of the integrin αvß3-FAK/Src pathway likely underlies vWF-induced endothelial dysfunction in sepsis. In conclusion, ISE protects against sepsis by inhibiting endothelial hyperpermeability and platelet-endothelium interactions.

USP36 inhibits apoptosis by deubiquitinating cIAP1 and survivin in colorectal cancer cells.

Chemotherapeutic agents for treating colorectal cancer (CRC) primarily induce apoptosis in tumor cells. The ubiquitin-proteasome system is critical for apoptosis regulation. Deubiquitinating enzymes (DUBs) remove ubiquitin from substrates to reverse ubiquitination. Although over 100 DUB members have been discovered, the biological functions of only a small proportion of DUBs have been characterized. Here, we aimed to systematically identify the DUBs that contribute to the development of CRC. Among the DUBs, ubiquitin-specific protease 36 (USP36) is upregulated in CRC. We showed that the knockdown of USP36 induces intrinsic and extrinsic apoptosis. Through gene silencing and coimmunoprecipitation techniques, we identified survivin and cIAP1 as USP36 targets. Mechanistically, USP36 binds and removes lysine-11-linked ubiquitin chains from cIAP1 and lysine-48-linked ubiquitin chains from survivin to abolish protein degradation. Overexpression of USP36 disrupts the formation of the XIAP-second mitochondria-derived activator of caspase complex and promotes receptor-interacting protein kinase 1 ubiquitination, validating USP36 as an inhibitor to intrinsic and extrinsic apoptosis through deubiquitinating survivin and cIAP1. Therefore, our results suggest that USP36 is involved in CRC progression and is a potential therapeutic target.

Multiple stresses induced by chronic exposure to flupyradifurone affect honey bee physiological states.

Flupyradifurone (FPF) has been reported to have a potential risk to terrestrial and aquatic ecosystems. In the present study, the effects of chronic FPF exposure on bees were systematically investigated at the individual behavioral, tissue, cell, enzyme activity, and the gene expression levels. Chronic exposure (14 d) to FPF led to reduced survival (12 mg/L), body weight gain (4 and 12 mg/L), and food utilization efficiency (4 and 12 mg/L). Additionally, FPF exposure (12 mg/L) impaired sucrose sensitivity and memory of bees. Morphological analysis revealed significant cellular and subcellular changes in brain neurons and midgut epithelial cells, including mitochondrial damage, nuclear disintegration, and apoptosis. FPF exposure (4 and 12 mg/L) led to oxidative stress, as evidenced by increased lipid peroxidation and alterations in antioxidant enzyme activity. Notably, gene expression analysis indicated significant dysregulation of apoptosis, immune, detoxification, sucrose responsiveness and memory-related genes, suggesting the involvement of different pathways in FPF-induced toxicity. The multiple stresses and potential mechanisms described here provide a basis for determining the intrinsic toxicity of FPF.

Discovery of anti-inflammatory agents from 3, 4-dihydronaphthalene-1(2H)-one derivatives by inhibiting NLRP3 inflammasome activation.

NLRP3 inflammatory vesicles are a polymer of cellular innate immunity composed of a pair of proteins. The continuous activation of NOD-like receptor pyrin domain-containing protein 3 (NLRP3) inflammatory vesicles induces the occurrence and enhancement of inflammatory response. In this study, a series of 3, 4-dihydronaphthalene-1(2H)-one derivatives (DHNs, 6a-u, 7a-e, 8a-n) were synthesized and characterized by NMR and HRMS. We evaluated the cytotoxicity and anti-inflammatory activity of all compounds in vitro, and selected 7a substituted by 7-Br in A-ring and 2-pyridylaldehyde in C-ring as effective lead compounds. Specifically, 7a can block the assembly and activation of NLRP3 inflammasome by down-regulating the expression of NLPR3 and apoptosis-associated speck-like protein containing a CARD (ASC), and inhibiting the production of reactive oxygen species (ROS) and other inflammatory mediators. In addition, 7a inhibits the phosphorylation of inhibitor kappa B alpha (IκBα) and NF-κB/p65 and the nuclear translocation of p65, thereby inhibiting nuclear factor kappa-B (NF-κB) signaling. Molecular docking analysis confirmed that 7a could reasonably bind the active sites of NLRP3, ASC and p65 proteins. Therefore, 7a is predicted as a potential NLRP3 inflammatory vesicle inhibitor and deserves further research and development.

p21 promotes gemcitabine tolerance in A549 cells by inhibiting DNA damage and altering the cell cycle.

Gemcitabine is one of the most widely used chemotherapy drugs for advanced malignant tumors, including non-small cell lung cancer. However, the clinical efficacy of gemcitabine is limited due to drug resistance. The aim of the present study was to investigate the role of p21 in gemcitabine-resistant A549 (A549/G+) lung cancer cells. IC50 values were determined using a Cell Counting Kit-8 (CCK-8) assay. mRNA and protein expression levels of genes were measured by reverse transcription-quantitative PCR and western blotting, respectively. The cell cycle distribution and apoptosis rate were analyzed by flow cytometry. DNA damage in cells was evaluated by single-cell gel electrophoresis. The results of western blot analysis and the CCK-8 assay demonstrated that the expression of p21 was higher in A549/G+ cells than in gemcitabine-sensitive cells. Knockdown of p21 expression in gemcitabine-resistant cells sensitized these cells to gemcitabine (with the IC50 decreasing from 84.2 to 26.7 µM). Cell cycle analysis revealed different changes in the cell cycle distribution in A549/G+ cells treated with the same concentration of gemcitabine, and decreased expression of p21 was shown to promote G1 arrest. The apoptosis assay and comet assay results revealed that decreased p21 expression resulted in accumulation of unrepaired DNA double-strand breaks (DSBs) and induction of apoptosis by gemcitabine. The present study demonstrated that knockout of p21 mRNA expression in A549/G+ cells promotes apoptosis and DNA DSB accumulation, accompanied by G1 arrest. These results indicated that p21 is involved in regulating the response of A549 cells to gemcitabine.

Unravelling and engineering an operon involved in the side-chain degradation of sterols in Mycolicibacterium neoaurum for the production of steroid synthons.

BACKGROUND: Harnessing engineered Mycolicibacteria to convert cheap phytosterols into valuable steroid synthons is a basic way in the industry for the production of steroid hormones. Thus, C-19 and C-22 steroids are the two main types of commercial synthons and the products of C17 side chain degradation of phytosterols. During the conversion process of sterols, C-19 and C-22 steroids are often produced together, although one may be the main product and the other a minor byproduct. This is a major drawback of the engineered Mycolicibacteria for industrial application, which could be attributed to the co-existence of androstene-4-ene-3,17-dione (AD) and 22-hydroxy-23,24-bisnorchol-4-ene-3-one (HBC) sub-pathways in the degradation of the sterol C17 side chain. Since the key mechanism underlying the HBC sub-pathway has not yet been clarified, the above shortcoming has not been resolved so far. RESULTS: The key gene involved in the putative HBC sub-pathway was excavated from the genome of M. neoaurum by comparative genomic analysis. Interestingly, an aldolase- encoding gene, atf1, was identified to be responsible for the first reaction of the HBC sub-pathway, and it exists as a conserved operon along with a DUF35-type gene chsH4, a reductase gene chsE6, and a transcriptional regulation gene kstR3 in the genome. Subsequently, atf1 and chsH4 were identified as the key genes involved in the HBC sub-pathway. Therefore, an updated strategy was proposed to develop engineered C-19 or C-22 steroid-producing strains by simultaneously modifying the AD and HBC sub-pathways. Taking the development of 4-HBC and 9-OHAD-producing strains as examples, the improved 4-HBC-producing strain achieved a 20.7 g/L production titer with a 92.5% molar yield and a 56.4% reduction in byproducts, and the improved 9-OHAD producing strain achieved a 19.87 g/L production titer with a 94.6% molar yield and a 43.7% reduction in byproduct production. CONCLUSIONS: The excellent performances of these strains demonstrated that the primary operon involved in the HBC sub-pathway improves the industrial strains in the conversion of phytosterols to steroid synthons.

Chronic larval exposure to thiacloprid impairs honeybee antennal selectivity, learning and memory performances.

The use of agricultural neonicotinoid insecticides has sub-lethal chronic effects on bees that are more prevalent than acute toxicity. Among these insecticides, thiacloprid, a commonly used compound with low toxicity, has attracted significant attention due to its potential impact on the olfactory and learning abilities of honeybees. The effect of sub-lethal larval exposure to thiacloprid on the antennal activity of adult honeybees (Apis mellifera L.) is not yet fully understood. To address this knowledge gap, laboratory-based experiments were conducted in which honeybee larvae were administered thiacloprid (0.5 mg/L and 1.0 mg/L). Using electroantennography (EAG), the impacts of thiacloprid exposure on the antennal selectivity to common floral volatiles were evaluated. Additionally, the effects of sub-lethal exposure on odor-related learning and memory were also assessed. The results of this study reveal, for the first time, that sub-lethal larval exposure to thiacloprid decreased honeybee antenna EAG responses to floral scents, leading to increased olfactory selectivity in the high-dose (1.0 mg/L) group compared to the control group (0 mg/L vs. 1.0 mg/L: p = 0.042). The results also suggest that thiacloprid negatively affected odor-associated paired learning acquisition, as well as medium-term (1 h) (0 mg/L vs. 1.0 mg/L: p = 0.019) and long-term memory (24 h) (0 mg/L vs. 1.0 mg/L: p = 0.037) in adult honeybees. EAG amplitudes were dramatically reduced following R-linalool paired olfactory training (0 mg/L vs. 1.0 mg/L: p = 0.001; 0 mg/L vs. 0.5 mg/L: p = 0.027), while antennal activities only differed significantly in the control between paired and unpaired groups. Our results indicated that exposure to sub-lethal concentrations of thiacloprid may affect olfactory perception and learning and memory behaviors in honeybees. These findings have important implications for the safe use of agrochemicals in the environment.

Reciprocal costimulatory molecules control the activation of mucosal type 3 innate lymphoid cells during engagement with B cells.

Innate lymphoid cells (ILCs) are the counterpart of T helper cells in the innate immune system and share multiple phenotypes with T helper cells. Inducible T-cell costimulator (ICOS) is recognized on T cells and participates in T-cell activation and T and B-cell engagement in lymphoid tissues. However, the role of ICOS in ILC3s and ILC3-involved interactions with the immune microenvironment remains unclear. Here, we found that ICOS expression on human ILC3s was correlated with the activated state of ILC3s. ICOS costimulation enhanced the survival, proliferation, and capacity of ILC3s to produce cytokines (IL-22, IL-17A, IFN-γ, TNF, and GM-CSF). Via synergistic effects of ICOS and CD40 signaling, B cells promoted ILC3 functions, and ILC3-induced T-cell-independent B-cell IgA and IgM secretion primarily required CD40 signaling. Hence, ICOS is essential for the nonredundant role of ILC3s and their interaction with adjacent B cells.

Thiacloprid impairs honeybee worker learning and memory with inducing neuronal apoptosis and downregulating memory-related genes.

Neonicotinoids are among the most widely used insecticides in the world and are recognized as a potential cause of pollinator decline. Previous studies have demonstrated that the neonicotinoid thiacloprid has adverse effects on foraging and memory behaviors. However, there is no direct evidence linking thiacloprid-induced neuronal cell damage in the brains of honeybees to learning and memory dysfunction. Adult honeybee (Apis mellifera L.) workers were chronically exposed to sub-lethal concentrations of thiacloprid. We discovered that thiacloprid negatively affected their survival, food consumption, and body weight. In addition, sucrose sensitivity and memory performance were impaired. We evaluated the apoptosis of honeybee brain cells using TUNEL (Terminal deoxynucleotidyl transferase-mediated digoxigenin-dUTP-biotin nick-end labeling) and Caspase-3 assays, which revealed that thiacloprid increases the dose-dependent apoptosis of neurons in the mushroom bodies (MB) and antennal lobes (AL). We also determined the abnormal transcripts of multiple genes, including vitellogenin (Vg), immune system genes (apidaecin and catalase), and memory-associated genes (pka, creb, Nmdar1, Dop2, Oa1, Oa-2R, and Oa-3R). These results indicate that exposure to sublethal concentrations of thiacloprid cause abnormal expression of memory-related genes and apoptosis of brain cells in the AL and MB, which may contribute to the memory disorder induced by thiacloprid exposure.

MiR-17- 5p/RRM2 regulated gemcitabine resistance in lung cancer A549 cells.

The main objective of this study is to investigate the regulatory roles of the miR-17-5p/RRM2 axis in A549/G+ cells' gemcitabine resistance. The cell viability was determined using CCK8 and clonogenic assays. Gene expression level analysis by RT-qPCR and Western blotting. Cell cycle analysis by flow cytometry. The dual luciferase activity assay was used to verify the target gene of miR-17-5p. In gemcitabine-resistant cell line A549G+, the drug resistance decreased after up-regulation of MiR-17-5p expression. The proportion of cell cycle G1 phase increased, and the S phase decreased. The expression level of cell cycle-related proteins CCNE1, CCNA2, and P21 decreased. The opposite results emerged after the down-regulation of MiR-17-5p expression in gemcitabine-sensitive cell line A549G-. The expression levels of PTEN and PIK3 in A549G+ cells were higher than in A549G-cells, but p-PTEN was lower than that in A549G-. After up-regulating the expression of MiR-17-5p in A549G+, the expression levels of p-PTEN increased, and the expression level of p-AKT decreased. After down-regulating miR-17-5p expression, the opposite results emerged. The dual-luciferase reporter assay and restorative experiments proved that RRM2 is one of the target genes for MiR-17-5p. Our results suggested that the miR-17-5p/RRM2 axis could adjust gemcitabine resistance in A549 cells, and the p-PTEN/PI3K/AKT signal pathway might be involved in this regulatory mechanism.

Distribution, sources, and risk assessment of polycyclic aromatic hydrocarbons (PAHs) in Kaokaowusu river sediments near a coal industrial zone.

This study systematically analyzed the contents, compositions, and sources of polycyclic aromatic hydrocarbons (PAHs) in river sediments near an important energy and chemical base in northwest China. In addition, their possible adverse effects on the ecology and human health were assessed. The PAH concentrations in this study area ranged from 2641.28 to 16783.72 (ng/g dw). PAHs of medium molecular weight (3-ring and 4-ring) showed the largest proportion, followed by PAHs of higher molecular weight (5-ring and 6-ring). The results of molecular diagnostic ratios and principal component analysis revealed that PAHs in the region have complex sources, with incomplete combustion of local fossil fuels and traffic exhaust factors being the main sources. The total toxic equivalent concentration of PAHs varied from 10.05 to 760.26 ng/g, and according to the sediment quality guidelines, PAHs have high potential ecological risk in the lower reaches of the river. The mean effect range-median quotient for the region was 0.46, and the combined ecological risk was at moderate to high levels (21% probability of toxicity). The lifetime carcinogenic risks for adults and children exposed to PAHs were 2.95 × 10-3 and 1.87 × 10-2, respectively, which are much higher than the limit of 10-4, indicating moderate to high potential cancer risks. Therefore, the local government should consider taking some environmental remediation measures. This study can provide theoretical support for pollution prevention measures and ecological restoration strategies for rivers in resource-rich areas.

Early-Life Sublethal Thiacloprid Exposure to Honey Bee Larvae: Enduring Effects on Adult Bee Cognitive Abilities.

Honey bees have significant ecological and economic value as important pollinators, but they are continuously exposed to various environmental stressors, including insecticides, which can impair their health and cause colony decline. (1) Background: Cognitive abilities are vital for the functional maintenance of honey bees; however, it remains unknown if chronic, low-dose exposure to thiacloprid during the larval stage impairs the cognitive abilities of emerged adult honey bees. (2) Methods: To explore this question, honey bee larvae were fed 0, 0.5, and 1.0 mg/L thiacloprid during their developmental phase. Then, the cognitive (i.e., olfactory learning and memory) abilities of adult honey bees were quantified to assess the delayed impacts of early-stage thiacloprid exposure on adult honey bee cognition. Neural apoptosis and transcriptomic level were also evaluated to explore the neurological mechanisms underlying these effects. (3) Results: Our results revealed that chronic larval exposure to sublethal thiacloprid impaired the learning and memory abilities of adult honey bees by inducing neuronal apoptosis and transcriptomic alterations. (4) Conclusions: We highlighted a previously unknown impairment caused by thiacloprid in honey bees.

Multiplexed site-specific genome engineering in Mycolicibacterium neoaurum by Att/Int system.

Genomic integration of genes and pathway-sized DNA cassettes is often an indispensable way to construct robust and productive microbial cell factories. For some uncommon microbial hosts, such as Mycolicibacterium and Mycobacterium species, however, it is a challenge. Here, we present a multiplexed integrase-assisted site-specific recombination (miSSR) method to precisely and iteratively integrate genes/pathways with controllable copies in the chromosomes of Mycolicibacteria for the purpose of developing cell factories. First, a single-step multi-copy integration method was established in M. neoaurum by a combination application of mycobacteriophage L5 integrase and two-step allelic exchange strategy, the efficiencies of which were ∼100% for no more than three-copy integration events and decreased sharply to ∼20% for five-copy integration events. Second, the R4, Bxb1 and ΦC31 bacteriophage Att/Int systems were selected to extend the available integration toolbox for multiplexed gene integration events. Third, a reconstructed mycolicibacterial Xer recombinases (Xer-cise) system was employed to recycle the selection marker of gene recombination to facilitate the iterative gene manipulation. As a proof of concept, the biosynthetic pathway of ergothioneine (EGT) in Mycolicibacterium neoaurum ATCC 25795 was achieved by remodeling its metabolic pathway with a miSSR system. With six copies of the biosynthetic gene clusters (BGCs) of EGT and pentose phosphate isomerase (PRT), the titer of EGT in the resulting strain in a 30 mL shake flask within 5 days was enhanced to 66 mg/L, which was 3.77 times of that in the wild strain. The improvements indicated that the miSSR system was an effective, flexible, and convenient tool to engineer the genomes of Mycolicibacteria as well as other strains in the Mycobacteriaceae due to their proximate evolutionary relationships.

A single gene integrates sex and hormone regulators into sexual attractiveness.

Sex differentiation and hormones are essential for the development of sexual signals in animals, and the regulation of sexual signals involves complex gene networks. However, it is unknown whether a core gene is able to connect the upstream regulators for controlling sexual signal outputs and behavioural consequences. Here, we identify a single gene that integrates both sex differentiation and hormone signalling with sexual attractiveness in an insect model. CYP4PC1 in the German cockroach, Blattella germanica, controls the rate-limiting step in producing female-specific contact sex pheromone (CSP) that stimulates male courtship. As revealed by behavioural, biochemical, molecular, genetic and bioinformatic approaches, in sexually mature females, CYP4PC1 expression and CSP production are coordinately induced by sex differentiation genes and juvenile hormone (JH) signalling. In adult males, direct inhibition of CYP4PC1 expression by doublesexM binding in gene promoter and lack of the gonadotropic hormone JH prevent CSP production, thus avoiding male-male attraction. By manipulating the upstream regulators, we show that wild-type males prefer to court cockroaches with higher CYP4PC1 expression and CSP production in a dose-dependent manner, regardless of their sex. These findings shed light on how sex-specific and high sexual attractiveness is conferred in insects.

Abnormal Antennal Olfactory Sensilla Phenotypes Involved in Olfactory Deficit in Bactrocera correcta (Diptera: Tephritidae).

The guava fruit fly, Bactrocera correcta, is one of the most destructive pests in the genus Bactrocera and detects environmental odorants mainly through antennal olfactory sensilla phenotypes with nanopores. However, it is unclear whether there are naturally occurring abnormal antennal olfactory sensilla phenotypes that affect olfaction. Here, we found that there were abnormal bulges besides nanopores on the surface of trichoid and basiconic olfactory sensilla in the antennal flagellum of long-term laboratory rearing colony (LTC), and that nanopore number in these olfactory sensilla was also remarkably reduced. Notably, the electroantennogram (EAG) responses of LTC insects to methyl eugenol or ß-caryophyllene were inhibited, and their behavioral responses elicited by the same odorants were also impaired. These results revealed naturally occurring abnormal antennal olfactory sensilla phenotypes which were involved in olfactory deficit in B. correcta, providing a platform to further study nanopore-targeted pest control technologies in the future.

One-pot biosynthesis of 7ß-hydroxyandrost-4-ene-3,17-dione from phytosterols by cofactor regeneration system in engineered mycolicibacterium neoaurum.

BACKGROUND: 7ß-hydroxylated steroids (7ß-OHSt) possess significant activities in anti-inflammatory and neuroprotection, and some of them have been widely used in clinics. However, the production of 7ß-OHSt is still a challenge due to the lack of cheap 7ß-hydroxy precursor and the difficulty in regio- and stereo-selectively hydroxylation at the inert C7 site of steroids in industry. The conversion of phytosterols by Mycolicibacterium species to the commercial precursor, androst-4-ene-3,17-dione (AD), is one of the basic ways to produce different steroids. This study presents a way to produce a basic 7ß-hydroxy precursor, 7ß-hydroxyandrost-4-ene-3,17-dione (7ß-OH-AD) in Mycolicibacterium, for 7ß-OHSt synthesis. RESULTS: A mutant of P450-BM3, mP450-BM3, was mutated and engineered into an AD producing strain for the efficient production of 7ß-OH-AD. The enzyme activity of mP450-BM3 was then increased by 1.38 times through protein engineering and the yield of 7ß-OH-AD was increased from 34.24 mg L- 1 to 66.25 mg L- 1. To further enhance the performance of 7ß-OH-AD producing strain, the regeneration of nicotinamide adenine dinucleotide phosphate (NADPH) for the activity of mP450-BM3-0 was optimized by introducing an NAD kinase (NADK) and a glucose-6-phosphate dehydrogenase (G6PDH). Finally, the engineered strain could produce 164.52 mg L- 1 7ß-OH-AD in the cofactor recycling and regeneration system. CONCLUSIONS: This was the first report on the one-pot biosynthesis of 7ß-OH-AD from the conversion of cheap phytosterols by an engineered microorganism, and the yield was significantly increased through the mutation of mP450-BM3 combined with overexpression of NADK and G6PDH. The present strategy may be developed as a basic industrial pathway for the commercial production of high value products from cheap raw materials.

Mesenchymal Stem Cells Alleviate Inflammatory Bowel Disease Via Tr1 Cells.

Mesenchymal stem cells (MSCs) have been used to achieve exciting therapeutic outcomes in many animal studies and clinical trials for various autoimmune diseases, including inflammatory bowel disease (IBD). Type 1 regulatory T (Tr1) cells are the main source of interleukin (IL) 10 in the intestine. Whether Tr1 cells are involved during MSC-mediated IBD treatment is unclear. We treated a murine model of 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis with human umbilical cord-derived MSCs (hUCMSCs) and found that the disease severity was alleviated significantly in a dose-dependent manner. hUCMSCs increased the proportion of Tr1 cells and decreased that of T helper (Th)-1 and Th17 cells in the spleen and mesenteric lymph nodes in different stages of colitis. We found that the upregulation of Tr1 cells by hUCMSCs was abrogated after blocking indoleamine-2,3-dioxygenase (IDO), and IDO knockdown in hUCMSCs reversed the increase in Tr1 cell proportions caused by hUCMSCs in colitis. Moreover, hUCMSCs inhibited apoptosis and promoted the proliferation of Tr1 cells. Our results suggest that Tr1 cells play an important role in the amelioration of IBD by MSCs, and they are the target population for the alleviation of IBD by MSCs, providing meaningful references for the study of therapeutic mechanisms of MSCs in other inflammatory diseases.

Commentary: How Do Microglia Regulate Neural Circuit Connectivity and Activity in the Adult Brain?

Microglia are the primary immune cells in CNS. Recent work shows that microglia are also essential for proper brain development through synaptic pruning and remodeling during early life development. But the question of whether and how microglia regulate synaptic connectivity in the adult brain remains open. Our recently published study provides new insights into the functional roles of microglia in the adult mouse brain. We find that chronic depletion of microglia via CSF1R inhibitors in the visual cortex in adult mice induces a dramatic increase in perineuronal nets, and enhances neural activities of both excitatory neurons and parvalbumin interneurons. These findings highlight new potential therapeutic avenues to enhance adult neural plasticity by manipulating microglia.

Role of DNA Flow Cytometry in the Diagnosis of Malignancy in Bile Duct Biopsies Using Paraffin-Embedded Tissue.

OBJECTIVES: Histopathologic evaluation of bile biopsies for biliary strictures is frequently challenging and is affected by interobserver disagreement. Reliable ancillary tests that can help differentiate benign from malignant are not available. This study aimed to evaluate whether DNA content abnormalities detected by flow cytometry on formalin-fixed, paraffin-embedded (FFPE) tissue can help differentiate benign/reactive, dysplastic from malignant cell populations in bile duct biopsies. METHODS: We performed DNA flow cytometry on 30 FFPE bile duct biopsies in 5 well-defined diagnostic categories: (1) negative for dysplasia (NED), (2) low-grade dysplasia (LGD), (3) high-grade dysplasia (HGD), (4) carcinoma (CA), and (5) indefinite for dysplasia (IND). RESULTS: Abnormal DNA content was detected in 0 NED, 5 LGD (62.5%), 2 HGD (33.3%), 3 CA (60%), and 4 IND (80%) samples. As a diagnostic marker, the estimated sensitivity, specificity, positive predictive value, and negative predictive value were 63%, 100%, 100%, and 50%, respectively, for diagnosing HGD or CA. CONCLUSIONS: DNA flow cytometry analysis is a useful ancillary test for the interpretation of bile duct biopsies. DNA content abnormalities, when correlated with histologic findings, will not only help confirm the morphologic impression but also identify patients who are at a higher risk of developing malignancy.