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Spider angioma refers to a type of telangiectasis that presents slightly beneath the skin surface on the face, neck, arms or upper trunk, often manifesting with a central red spot and reddish extensions that radiate outwards like a spider's web. The cutaneous spider angioma may be benign but it often indicates abnormal liver function, especially in patients with chronic cirrhosis. The spider angioma is irreversible and rarely occurred diffusely over the body or with giant sizes. Here, we report two rare multiple and giant spider angioma cases in patients with HIV/AIDS who developed chronic cirrhosis. In addition, we comprehensively reviewed related literatures and evaluated the existing possible mechanisms of spider angioma.

HMOs (human milk oligosaccharides) are the third most important nutrient in breast milk. As complex glycans, HMOs play an important role in regulating neonatal intestinal immunity, resisting viral and bacterial infections, displaying anti-inflammatory characteristics, and promoting brain development. Although there have been some previous reports of HMOs, a detailed literature review summarizing the structure-activity relationships and dose-dependent effects of HMOs is lacking. Hence, after introducing the structures and synthetic pathways of HMOs, this review summarizes and categorizes identified structure-function relationships of HMOs. Differential mechanisms of different structural HMOs utilization by microorganisms are summarized. This review also emphasizes the recent advances in the interactions between different health benefits and the variance of dosage effect based on in vitro cell tests, animal experiments, and human intervention studies. The potential relationships between the chemical structure, the dosage selection, and the physiological properties of HMOs as functional foods are vital for further understanding of HMOs and their future applications.

Many species of devastating insect pests have acquired a high degree of resistance to insecticides in the field during the last few decades. Spodoptera exigua, for example, is the most damaging pests of economic crops with a worldwide spread. In a present study, the comparative growth, reproduction, and detoxification enzyme activity were evaluated along with exposure to three insecticides at low lethal doses of lufenuron, indoxacarb, and spinosad as compared to the control. Results indicate that the larval developmental time was significantly extended on lufenuron (21.5 ± 29 days) followed by indoxacarb (20.28 ± 0.24 days) and spinosad (19.74 ± 0.23 days) as compared to that on the control (18.13 ± 0.13 days). Similarly, the lowest number of eggs of S. exigua females were recorded on lufenuron (328.75 ± 50.81 eggs) followed by spinosad (367 ± 36.4 eggs) and indoxacarb (411.58 ± 42.38 eggs) as compared to that on the control (560.2 ± 13.47). Interestingly, the lowest intrinsic rate of increase (r) (0.121 ± 0.009) and highest mean generation time (T) (36.2 ± 0.35 days) were observed when larvae were treated to a low lethal concentration (LC

Understanding how the tumor microenvironment is shaped by various factors is important for the development of new therapeutic strategies. Tumor cells often undergo spontaneous apoptotic cell death in tumor microenvironment, these apoptotic cells are histologically co-localized with immunosuppressive macrophages. However, the mechanism by which tumor cell apoptosis modulates macrophage polarization is not fully understood. In this study, we aimed to explore the tumor promoting effects of apoptotic tumor cells and the signal pathways involved.Apoptotic cells and macrophages in tumors were detected by immunohistochemical staining. Morphological analysis was performed with Giemsa staining. Lipids generated from apoptotic cells were detected by liquid chromatography-mass spectrometry. Phosphatidylserine-containing liposomes were prepared to mimic apoptotic cells. The expression of protein was determined by real-time PCR, immunohistochemistry enzyme-linked immunosorbent assay and Western blotting. Mouse malignant ascites and subcutaneous tumor models were designed for in vivo analysis. Transgenic mice with specific genes knocked out and inhibitors specific to certain proteins were used for the mechanistic studies.The location and the number of apoptotic cells were correlated with that of macrophages in several types of carcinomas. Phosphatidylserine, a lipid molecule generated in apoptotic cells, induced polarization and accumulation of M2-like macrophages in vivo and in vitro. Moreover, sustained administration of phosphoserine promoted tumor growth in the malignant ascites and subcutaneous tumor models. Further analyses suggested that phosphoserine induced a M2-like phenotype in macrophages, which was related to the activation of phosphoserine receptors including T-cell immunoglobin mucin 4 (TIM4) and the FAK-SRC-STAT3 signaling pathway as well as elevated the expression of the histone demethylase Jumonji domain-containing protein 3 (JMJD3). Administration of specific inhibitors of these pathways could reduce tumor progression.This study suggest that apoptotic cell-generated phosphoserine might be a notable signal for immunosuppressive macrophages in tumors, and the related pathways might be potential therapeutic targets for cancer therapy.

Cholestasis is a major cause of a series of bile flow malfunction-related liver diseases. Pregnane X receptor (PXR) is a key regulator in endo- and xeno-biotics metabolism, which has been considered as a promising therapeutic target for cholestasis. In this study we conducted human PXR (hPXR) agonistic screening using dual-luciferase reporter gene assays, which led to discovering a series of potent hPXR agonists from a small Euphorbiaceae diterpenoid library, containing 35 structurally diverse diterpenoids with eight different skeleton types. The most active compound 6, a lathyrane diterpenoid (5/11/3 ring system), dose-dependently activated hPXR with a high selectivity, and significantly upregulated the expression of hPXR downstream genes CYP3A4 and UGT1A1. In LCA-induced cholestasis mouse model, administration of compound 6 (50 mg· kg

Calreticulin (Crt) is a highly conserved and multi-functional protein with lectin-like properties and important immunological activities. In this study, a Crt homolog, namely, ToCrt, was cloned and characterized from the obscure puffer Takifugu obscurus with an open reading frame of 1278 bp encoding a putative protein of 425 amino acids. The deduced amino acid sequence of ToCrt consisted of three conserved structural domains: N-domain, P-domain, and C-terminal domain. In the phylogenetic tree, ToCrt formed a separate cluster with three Crts from other pufferfish species (Takifugu rubripes, Takifugu flavidus, and Takifugu bimaculatus). The mRNA transcript of ToCrt was ubiquitously expressed in all the examined tissues in a decreasing order: liver, spleen, kidney, gills, intestine, and heart. After Vibrio harveyi, Edwardsiella tarda, and Aeromonas hydrophila stimulations, the levels of ToCrt mRNA in the kidney and spleen were significantly upregulated compared with that in the control group. The recombinant calreticulin domain of ToCrt (rToCrt) could bind three Gram-negative bacteria (V. harveyi, E. tarda, and A. hydrophila) and polysaccharides from bacterial cell walls such as lipopolysaccharide and peptidoglycan. Meanwhile, rToCrt could agglutinate different kinds of microorganisms and exhibit antimicrobial activity. These results suggested that T. obscurus ToCrt could serve as an antimicrobial effector in the host immune response against invading microorganisms.

To observe the effect of electroacupuncture (EA) on behavior, pathomorphology and brain cell apoptosis in traumatic brain injury (TBI) rats, so as to explore its mechanisms underlying treatment of TBI.Male SD rats were randomly divided into control (After modeling, the scores of balance and walking in the model group were higher than those in the sham group (EA can improve the behavioral function, reduce the apoptosis of brain cells in the injured area, and promote the rehabilitation of craniocerebral injury.

Objective Polycystic ovary syndrome (PCOS) is a common gynecological endocrine disease in reproductive women, and the endocrine levels are also affected by diseases. The aim of this study was to determine the effect of thrombospondin-1 (TSP-1) on PCOS rat model. Methods We established the PCOS rat model, the serum hormones including TSP-1 expression were determined and morphological characteristics were investigated to evaluate the model. These above endocrine and morphological features were investigated again to evaluate the effect of TSP-1 treatment. Results In the PCOS model group, the serum hormones change (higher luteinizing hormone, testosterone and estrogen) and decreased TSP-1 expression levels were found compared with the control group. Besides, the morphological characteristics of PCOS were also observed in the model group. After TSP-1 treatment, the higher TSP-1, ANGPT2, PDGFB and PDGFD expression levels, the lower LH and T levels, decreased vessel density as well as VEGFA and ANGPT1 expression levels were found compared with the control group, and the ovary morphological changes were also observed in the TSP-1 experimental group. Conclusions TSP-1 delivery system might be an alternative therapy for PCOS treatment.

BackgroundDespite the use of aggressive multimodality treatment, most anaplastic thyroid carcinoma (ATC) patients die within a year of diagnosis. Although the combination of BRAF and MEK inhibitors has recently been approved for use in BRAF-mutated ATC, they remain effective in a minority of patients who are likely to develop drug resistance. There remains a critical clinical need for effective systemic agents for ATC with a reasonable toxicity profile to allow for rapid translational development.Material and MethodsTwelve human thyroid cancer cell lines with comprehensive genomic characterization were used in a high-throughput screening (HTS) of 257 compounds to select agents with maximal growth inhibition. Cell proliferation, colony formation, orthotopic thyroid models, and patient-derived xenograft (PDX) models were used to validate the selected agents.ResultsSeventeen compounds were effective, and docetaxel, LBH-589, and pralatrexate were selected for additional in vitro and in vivo analysis as they have been previously approved by the US Food and Drug Administration for other cancers. Significant tumor growth inhibition (TGI) was detected in all tested models treated with LBH-589; pralatrexate demonstrated significant TGI in the orthotopic papillary thyroid carcinoma model and 2 PDX models; and docetaxel demonstrated significant TGI only in the context of mutant TP53.ConclusionsHTS identified classes of systemic agents that demonstrate preferential effectiveness against aggressive thyroid cancers, particularly those with mutant TP53. Preclinical validation in both orthotopic and PDX models, which are accurate in vivo models mimicking tumor microenvironment, may support initiation of early-phase clinical trials in non-BRAF mutated or refractory to BRAF/MEK inhibition ATC.

Context Ginsenoside Rb1, the main active ingredient of ginseng, exhibits ex vivo depression of store-operated calcium entry (SOCE) and related vasoconstriction in pulmonary arteries derived from pulmonary hypertension (PH) rats. However, the in vivo effects of ginsenoside Rb1 on PH remain unclear. Objective This study explored the possibility of using ginsenoside Rb1 as an in vivo preventive medication for type I PH, i.e., pulmonary arterial hypertension (PAH), and potential mechanisms involving SOCE. Materials and methods Male Sprague-Dawley rats (170–180 g) were randomly divided into Control, MCT, and MCT + Rb1 groups (n = 20). Control rats received only saline injection. Rats in the MCT + Rb1 and MCT groups were intraperitoneally administered single doses of 50 mg/kg monocrotaline (MCT) combined with 30 mg/kg/day ginsenoside Rb1 or equivalent volumes of saline for 21 consecutive days. Subsequently, comprehensive parameters related to SOCE, vascular tone, histological changes and hemodynamics were measured. Results Ginsenoside Rb1 reduced MCT-induced STIM1, TRPC1, and TRPC4 expression by 35.00, 31.96, and 32.24%, respectively, at the protein level. SOCE-related calcium entry and pulmonary artery contraction decreased by 162.6 nM and 71.72%. The mean pulmonary artery pressure, right ventricle systolic pressure, and right ventricular mass index decreased by 19.5 mmHg, 21.6 mmHg, and 39.50%. The wall thickness/radius ratios decreased by 14.67 and 17.65%, and the lumen area/total area ratios increased by 18.55 and 15.60% in intrapulmonary vessels with 51–100 and 101–150 μm o.d. Conclusion Ginsenoside Rb1, a promising candidate for PH prevention, inhibited SOCE and related pulmonary vasoconstriction, and relieved MCT-induced PAH in rats.

The microbiota of mites is closely related to their growth, development, and pathogenicity. Therefore, it is necessary to study the bacteria in mites. Here, for the first time, based on 16s rRNA V3-V4 region, the microbiota of 45 samples of nine species in six families of medically important mites were analyzed using Illumina MiSeq sequencing technique. The results showed that, at the phylum level, Proteobacteria (56.20–86.40%) were the dominant, followed by Firmicutes (6.41–19.43%), Bacteroidetes (5.56–13.38%) and Actinobacteria (1.93–28.07%). But at the genera the microbiota of mites are different, showing four characteristics: 1) The microbiota is related to the parasitic host. Demodex folliculorum (Acariforms: Demodicidae) and D. brevis (Acariforms: Demodicidae), both parasitizing humans, showed similar microbial composition, as did D. canis (Acariforms: Demodicidae) and Sarcoptes scabiei canis (Acariforms: Sarcoptidae) parasitizing dogs, but D. caprae (Acariforms: Demodicidae) parasitizing sheep showed unique microbial community; 2) The microbiota is related to mite’s species. Dermatophagoides farinae and Cheyletus malaccensis (Acariforms: Cheyletidae), both collecting from flour, show respective microbial composition; 3) The microbiota is related to the life stage. There were differences in microbiota between adults and larvae of D. farinae, but no differences observed in Psoroptes cuniculi (Acariforms: Psoroptidae); and 4) The microbiota is related to the blood-feeding state. The microbiota of blood-fed Ornithonyssus bacoti (Parasitiformes: Macronyssidae) adults was significantly higher than that of unfed adults. This indicates that the microbiota of mites is affected by mite species, parasitic host, growth stage and habitat. Therefore, understanding these influencing factors will have a very important guiding significance for the prevention and control of mite-borne diseases.

Gemcitabine (Gem) is currently used as the first-line therapy for liver and pancreatic cancer but has limited efficacy in most cases. Dexamethasone (Dex) have been applied as a chemoprotectant and chemosensitizer in cancer chemotherapy. This study further explored the potential of combination of Gem and Dex and tested the hypothesis that glucocorticoid receptor signaling is essential for the synergistic antitumor activity. In the HepG2 and AsPC-1 xenograft models, the combination treatment showed a significantly synergistic antitumor activity. Immunohistochemistry of post-treatment tumors showed a significant decrease in proliferation and angiogenesis as compared to either of the treatments alone. Dex alone and the combination with Gem inhibited the expression of glucocorticoid receptor. The combination of Dex and Gem showed synergistic cytotoxicity in cell lines in vitro. The antiproliferative synergism is prevented by used glucocorticoid receptor (GR) small interfering RNA, demonstrating that the glucocorticoid receptor is required for the antiproliferative synergism of Gem and Dex. The inhibition of glucocorticoid receptor signaling pathway and induction of apoptosis via activation of caspases 3, 8 and 9, PARP, contributed to the synergistic effect of this combination therapy. These results demonstrate that Dex could potentiate the antitumor efficacy of Gem. The synergistic antitumor activity of the combination of Dex and Gem was through glucocorticoid receptor signaling. Taken together, a combination of Dex and Gem shows a significant synergistic antitumor activity and lesser toxicity both in vitro and in vivo and could be a combination chemotherapy for the treatment of highly expression of glucocorticoid receptor patients.

Trimethylamine-N-oxide (TMAO) is a gut microbial metabolite that promotes Alzheimer’s disease (AD) progression. Given that probiotics can alleviate AD symptoms by inhibiting the synthesis of TMAO, here we investigated the correlation between TMAO and cognitive deterioration by measuring TMAO levels in the plasma of choline-treated APP/PS1 mice (an AD mouse model) with and without probiotic treatments. We found that declines in L. plantarum in the gut were associated with cognitive impairment. Moreover, 12-weeks of treatment with memantine plus L. plantarum ameliorated cognitive deterioration, decreased Αβ levels in the hippocampus, and protected neuronal integrity and plasticity. These effects were accompanied by reductions in TMAO synthesis and neuroinflammation. These experiments demonstrate that L. plantarum augments the beneficial therapeutic effects of memantine treatment in APP/PS1 mice by remodeling the intestinal microbiota, inhibiting the synthesis of TMAO, and reducing clusterin levels. Our results thus highlight intestinal microbiota as a potential therapeutic target to decrease the risk of AD.

Rationale: Pulmonary hypertension (PH) is a life-threatening cardiopulmonary disorder in which inflammation and immunity have emerged as critical early pathogenic elements. Although proinflammatory processes in PH and pulmonary arterial hypertension (PAH) are the focus of extensive investigation, the initiating mechanisms remain elusive. Objectives: We tested whether activation of the complement cascade is critical in regulating proinflammatory and pro-proliferative processes in the initiation of experimental hypoxic PH and can serve as a prognostic biomarker of outcome in human PAH. Methods: We used immunostaining of lung tissues from experimental PH models and patients with PAH, analyses of genetic murine models lacking specific complement components or circulating immunoglobulins, cultured human pulmonary adventitial fibroblasts, and network medicine analysis of a biomarker risk panel from plasma of patients with PAH. Measurements and Main Results: Pulmonary perivascular-specific activation of the complement cascade was identified as a consistent critical determinant of PH and PAH in experimental animal models and humans. In experimental hypoxic PH, proinflammatory and pro-proliferative responses were dependent on complement (alternative pathway and component 5), and immunoglobulins, particularly IgG, were critical for activation of the complement cascade. We identified Csf2/GM-CSF as a primary complement-dependent inflammatory mediator. Furthermore, using network medicine analysis of a biomarker risk panel from plasma of patients with PAH, we demonstrated that complement signaling can serve as a prognostic factor for clinical outcome in PAH. Conclusions: This study establishes immunoglobulin-driven dysregulated complement activation as a critical pathobiological mechanism regulating proinflammatory and pro-proliferative processes in the initiation of experimental hypoxic PH and demonstrates complement signaling as a critical determinant of clinical outcome in PAH.

Environmental DNA (eDNA) integrated with metabarcoding is a promising and powerful tool for species composition and biodiversity assessment in aquatic ecosystems and is increasingly applied to evaluate fish diversity. To date, however, no standardized eDNA-based protocol has been established to monitor fish diversity. In this study, we investigated and compared two filtration methods and three DNA extraction methods using three filtration water volumes to determine a suitable approach for eDNA-based fish diversity monitoring in the Pearl River Estuary (PRE), a highly anthropogenically disturbed estuarine ecosystem. Compared to filtration-based precipitation, direct filtration was a more suitable method for eDNA metabarcoding in the PRE. The combined use of DNeasy Blood and Tissue Kit (BT) and traditional phenol/chloroform (PC) extraction produced higher DNA yields, amplicon sequence variants (ASVs), and Shannon diversity indices, and generated more homogeneous and consistent community composition among replicates. Compared to the other combined protocols, the PC and BT methods obtained better species detection, higher fish diversity, and greater consistency for the filtration water volumes of 1 000 and 2 000 mL, respectively. All eDNA metabarcoding protocols were more sensitive than bottom trawling in the PRE fish surveys and combining two techniques yielded greater taxonomic diversity. Furthermore, combining traditional methods with eDNA analysis enhanced accuracy. These results indicate that methodological decisions related to eDNA metabarcoding should be made with caution for fish community monitoring in estuarine ecosystems.环境DNA（eDNA）宏条形码技术是评估水生生态系统中物种组成和生物多样性的强有力工具，并已广泛应用于鱼类多样性的监测。然而，目前，用于监测鱼类多样性的eDNA技术尚未标准化。该研究分别比较了两种过滤方法、三种过滤水样体积以及三种DNA提取方法的DNA产出效果，以期为珠江口等人为干扰频繁的河口生态系统提供一种合适的基于eDNA的鱼类多样性监测方法。结果表明，与基于过滤的沉淀法相比，直接过滤法更适用于珠江口的eDNA宏条形码技术研究；DNeasy Blood and Tissue Kit（BT）和苯酚/氯仿（PC）提取方法具有较高的DNA产量、扩增子序列变体（ASV）丰度和香农多样性指数，并可以获得更均匀和一致的鱼类群落组成；其中，PC和BT分别在提取过滤水样体积为1000 mL和2000 mL时，获得更高的物种检测率、鱼类多样性和种类组成一致性。与底拖网相比，eDNA宏条形码技术在珠江口鱼类多样性调查中具有更高的灵敏度，监测到的种类数量更多，但两种方法在种类组成上具有一定的差异，二者的结合有利于提高鱼类多样性监测的准确性。这些结果表明，在河口生态系统的鱼类群落监测中，eDNA宏条形码技术流程中相关步骤的方法选择需要慎重考虑。.

Uncontrolled inflammation causes health problems. Extracellular signal-regulated kinase (ERK) phosphorylates signal transducer and activator of transcription 3 (STAT3) at Ser727, resulting in inflammation. The leaf of Vernonia amygdalina (VA) is a medicinal herb for managing inflammation-associated diseases. Oral administration or topical application of VA leaf extract exerts anti-inflammatory effects in rat models. However, the anti-inflammatory mechanisms of the herb are not fully understood.In this study, we aimed to investigate the involvement of ERK/STAT3 (Ser727) signaling in the anti-inflammatory effects of an ethanolic extract of VA leaves.Extracts of VA leaves were prepared with different concentrations of ethanol. A LPS-stimulated RAW264.7 cell model was used for in vitro assays, and a TPA (12-O-tetradecanoylphorbol-13-acetate)-induced ear edema mouse model was employed for in vivo assays. The 95% ethanol extract of VA leaves (VAE) exerted the strongest inhibitory effect on nitric oxide (NO) production in LPS-stimulated macrophages; thus it was selected for use in this study. Hematoxylin and eosin (HE) staining was used to examine pathological conditions of mouse ear tissues. Griess reagent was employed to examine NO generation in cell cultures. Immunoblotting and ELISA were used to examine protein levels, and RT-qPCR was employed to examine mRNA levels.Topical application of VAE ameliorated mouse ear edema induced by TPA. VAE suppressed the phosphorylation of ERK (Thr202/Tyr204) and STAT3 (Ser727); and decreased protein levels of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), interleukin (IL)-6, IL-1β and tumor necrosis factor-α (TNF-α) in the mouse ear tissues and in LPS-stimulated RAW 264.7 cells. VAE also inhibited NO production, and lowered mRNA levels of IL-6, IL-1β and TNF-α in the macrophages.VAE ameliorates TPA-induced mouse ear edema. Suppression of ERK/STAT3 (Ser727) signaling is involved in VAE's anti-inflammatory effects. These novel data provide further pharmacological justifications for the medicinal use of VA in treating inflammation-associated diseases, and lay the groundwork for developing VAE into a new anti-inflammatory agent.

Lysozyme (Lyz) is an alkaline enzyme that hydrolyzes mucopolysaccharides in bacteria and is highly conserved vertebrates and invertebrates. In this study, a c-type lysozyme gene (named ToLyzC) from the obscure puffer Takifugu obscurus was cloned and characterized. The full-length cDNA of ToLyzC was 432 bp, encoding 143 amino acids, with a predicted molecular mass of 16.2 kDa and a theoretical pI of 8.86. The depicted protein sequence contained a LYZ1 domain from 16 to 142 amino acids, seven conserved cysteine residues. Phylogenetic analysis indicated that ToLyzC clustered with Lyzs from other teleost fishes. Quantitative real-time PCR analysis revealed that ToLyzC mRNA was mainly expressed in the liver. The transcript level of ToLyzC gene was significantly upregulated after Staphylococcus aureus and Vibrio harveyi challenge. The optimal pH and temperature of recombinant ToLyzC protein (rToLyzC) lytic activity was detected to be 7.5 and 35 °C, respectively. rToLyzC exhibited significant antibacterial and bacterial binding activities against S. aureus, Aeromonas hydrophila, V. harveyi, and Edwardsiella tarda at different time points. In addition, the morphological changes of V. harveyi cells treated with rToLyzC were observed under scanning electron microscope, which further confirmed the antibacterial and bacteriolytic activity of rToLyzC. Taken together, our current study indicated that ToLyzC is involved in the immune response to bacterial infection in obscure puffers.

Chicken manure is a source of antibiotic resistance genes (ARGs) and pathogenic microbes. Mikania micrantha Kunth (MM) is an invasive plant containing phytochemicals as antimicrobial agents. To explore its impacts on ARGs and pathogen-host interactions (PHIs), MM was added to composting mixtures. The findings indicated that compared with control (CK), MM significantly improved the phytochemical abundances, particularly stilbenoids and diarylheptanoids (4.87%), and ubiquinones (2.66%) in the treatment (T) compost. Besides, significant ARGs reduction was noted, where rpoB2, RbpA, FosB1, vatC, and vatB were removed from T compost. PHIs significantly declined in T compost, where the growth of Xanthomonas citri, Streptococcus pneumoniae, Fusarium graminearum, Vibrio cholerae, and Xanthomonas campestris were inhibited. Multiple variable analyses demonstrated that temperature and pH revealed a significant role in ARGs and PHIs decline. Accordingly, this study considerably recommends MM as a promising compost additive in terms of its antimicrobial potential toward pathogenic microbes and ARGs.

The lack of effective pharmacotherapies for nonalcoholic fatty liver disease (NAFLD) is mainly attributed to insufficient research on its pathogenesis. The pathogenesis of TM6SF2-efficient NAFLD remains unclear, resulting in a lack of therapeutic strategies for TM6SF2-deficient patients.To investigate the role of TM6SF2 in fatty acid metabolism in the context of fatty liver and propose possible therapeutic strategies for NAFLD caused by TM6SF2 deficiency.Liver samples collected from both NAFLD mouse models and human participants (80 cases) were used to evaluate the expression of TM6SF2 by using western blotting, immunohistochemistry, and quantitative polymerase chain reaction. RNA-seq data retrieved from the Gene Expression Omnibus database were used to confirm the over-expression of TM6SF2. Knockdown and overexpression of TM6SF2 were performed to clarify the mechanistic basis of hepatic lipid accumulation in NAFLD. MK-4074 administration was used as a therapeutic intervention to evaluate its effect on NAFLD caused by TM6SF2 deficiency.Hepatic TM6SF2 levels were elevated in patients with NAFLD and NAFLD mouse models. TM6SF2 overexpression can reduce hepatic lipid accumulation, suggesting a protective role for TM6SF2 in a high-fat diet (HFD). Downregulation of TM6SF2, simulating the TM6SF2 E167K mutation condition, increases intracellular lipid deposition due to dysregulated fatty acid metabolism and is characterized by enhanced fatty acid uptake and synthesis, accompanied by impaired fatty acid oxidation. Owing to the potential effect of TM6SF2 deficiency on lipid metabolism, the application of an acetyl-CoA carboxylase inhibitor (MK-4074) could reverse the NAFLD phenotypes caused by TM6SF2 deficiency.TM6SF2 plays a protective role in the HFD condition; its deficiency enhanced hepatic lipid accumulation through dysregulated fatty acid metabolism, and MK-4074 treatment could alleviate the NAFLD phenotypes caused by TM6SF2 deficiency.

Extracellular vesicles are cell-to-cell communication tools that play important roles in sperm maturation, motility, and fertilization. Epididymosomes provide proteins, lipids, and genetic materials to sperm as the latter go through the epididymis, not only promoting sperm maturation, but also influencing sperm and offspring phenotypes at the epigenetic level. After ejaculation, prostasomes fuse with sperm, leading to changes in the composition of the sperm plasma membrane and the contents of sperm, which maintains the stability of the sperm plasma membrane and increases sperm motility as well. Uterosomes can enhance sperm capacitation and acrosome reaction. Oviductosomes participate in sperm capacitation and motility, provide non-coding RNA to sperm, and influence the development of embryos. However, studies on extracellular vesicles and spermatozoa are mostly based on animal experiments, and their applicability to humans remains to be further explored.

Mitochondria play an essential role in pathophysiology of both inflammatory and neuropathic pain (NP), but the mechanisms are not yet clear. Dynamin-related protein 1 (Drp1) is broadly expressed in the central nervous system and plays a role in the induction of mitochondrial fission process. Spared nerve injury (SNI), due to the dysfunction of the neurons within the spinal dorsal horn (SDH), is the most common NP model. We explored the neuroprotective role of Drp1 within SDH in SNI. SNI mice showed pain behavior and anxiety-like behavior, which was associated with elevation of Drp1, as well as increased density of mitochondria in SDH. Ultrastructural analysis showed SNI induced damaged mitochondria into smaller perimeter and area, tending to be circular. Characteristics of vacuole in the mitochondria further showed SNI induced the increased number of vacuole, widened vac-perimeter and vac-area. Stable overexpression of Drp1 via AAV under the control of the Drp1 promoter by intraspinal injection (Drp1 OE) attenuated abnormal gait and alleviated pain hypersensitivity of SNI mice. Mitochondrial ultrastructure analysis showed that the increased density of mitochondria induced by SNI was recovered by Drp1 OE which, however, did not change mitochondrial morphology and vacuole parameters within SDH. Contrary to Drp1 OE, down-regulation of Drp1 in the SDH by AAV-Drp1 shRNA (Drp1 RNAi) did not alter painful behavior induced by SNI. Ultrastructural analysis showed the treatment by combination of SNI and Drp1 RNAi (SNI + Drp1 RNAi) amplified the damages of mitochondria with the decreased distribution density, increased perimeter and area, as well as larger circularity tending to be more circular. Vacuole data showed SNI + Drp1 RNAi increased vacuole density, perimeter and area within the SDH mitochondria. Our results illustrate that mitochondria within the SDH are sensitive to NP, and targeted mitochondrial Drp1 overexpression attenuates pain hypersensitivity. Drp1 offers a novel therapeutic target for pain treatment., Graphical abstract Image 1, Highlights • SNI induces pain hypersensitivity and increases Drp1 within SDH in mice. • Targeted up-regulation of Drp1 in SDH attenuates SNI-pain hypersensitivity. • Targeted up-regulation of Drp1 restores mitochondrial fragmentation and swelling. • Targeted down-regulation of Drp1 does not alleviate pain with the deteriorated mitochondrial damages. • Drp1 offers a novel therapeutic target for pain treatment.

Sirtuin-1 (SIRT1) is involved in various metabolic pathways, including fatty acid synthesis and gluconeogenesis in the liver. However, its role in initiation and progression of liver cancer remains unclear. Studying Sirt1 liver-specific knockout (LKO) mice in combination with diethylnitrosamine (DEN) treatment, we demonstrated that loss of Sirt1 rendered mice resistant to DEN-induced hepatocellular carcinoma (HCC) development. RNA-seq revealed that livers from LKO mice exhibited an enrichment in glutathione metabolism eight months after DEN challenge. Sirt1 deficiency elevated the expression of glutathione-s-transferase family genes by increasing the level of Nrf2, a key regulator of glutathione metabolism. Hence, LKO livers displayed a reductive environment with an increased ratio of GSH to GSSG and an elevated GSH level. Furthermore, using CRISPR knockout techniques, we confirmed that the impairment of HCC formation in LKO mice is mainly dependent on NRF2 signaling. Meanwhile, HCC induced by DEN could be blocked by the administration of N-acetyl cysteine (NAC) when administered one month after DEN challenge. However, NAC treatment starting five months after DEN injection was not able to prevent tumor development. In conclusion, our findings indicate that a reductive environment orchestrated by glutathione metabolism at an early stage can prevent the initiation of HCC.

Calnexin (Cnx) is a membrane-bound lectin chaperone of the endoplasmic reticulum. In this study, a novel Cnx homologue from the obscure puffer Takifugu obscurus was characterized, tentatively named ToCnx. The cDNA of ToCnx was 1803 bp, and it contained an open reading frame encoding a polypeptide of 600 amino acid residues with a calculated molecular weight of 67.5 kDa. Multiple alignment of the deduced amino acid sequences of ToCnx and other related fish Cnxs revealed that ToCnx had typical characteristics of fish Cnxs. Sequence comparison and phylogenetic tree analysis showed that ToCnx had the closest relationship with Cnxs from Takifugu flavidus and Takifugu rubripes. ToCnx transcripts were detected in all the tissues examined, and they were mainly expressed in the liver, kidney, and intestine. Upon Vibrio harveyi, Edwardsiella tarda, and Aeromonas hydrophila infection, ToCnx transcripts were all significantly upregulated in the kidneys. The recombinant calreticulin domain of ToCnx (rToCnx) was prepared by prokaryotic expression. In the absence of calcium, rToCnx was able to bind three Gram-negative bacteria (V. harveyi, E. tarda, and A. hydrophila) and two bacterial saccharides, such as lipopolysaccharide and peptidoglycan. In the presence of calcium, rToCnx could agglutinate all the detected microorganisms. In addition, rToCnx possessed the effect of inhibiting the growth of three microbe strains. These observations suggested that ToCnx is an important participant in host immune defense against bacteria.

Background Adventitial remodeling is a pathological hallmark of hypertension that results in target organ damage. Activated adventitial fibroblasts have emerged as critical regulators in this process, but the precise mechanism remains unclear. Methods and Results Interleukin 11 (IL‐11) knockout and wild‐type mice were subjected to angiotensin II (Ang II) infusion to establish models of hypertension‐associated vascular remodeling. IL‐11 mRNA and protein were increased especially in the adventitia in response to Ang II. Compared with wild‐type mice, Ang II–treated IL‐11 knockout mice showed amelioration of vascular hypertrophy, adventitial fibrosis, macrophage infiltration, and inflammatory factor expression. Recombination mouse IL‐11 exacerbated adventitial fibrosis in Ang II–infused wild‐type mice. Interestingly, IL‐11 neutralizing antibody attenuated adventitial fibrosis, macrophage infiltration, and inflammatory factor expression after Ang II infusion for 7 days. Mechanistically, in primary cultured adventitial fibroblasts, Krüppel‐like factor 15 negatively regulated Ang II–induced IL‐11 expression. Ang II increased extracellular signal‐regulated kinases 1 and 2 activation, especially in adventitia, and caused biphasic extracellular signal‐regulated kinases 1 and 2 activation in adventitial fibroblasts. A rapid and early activation increased IL‐11 production through decreasing Krüppel‐like factor 15 expression, which, in turn, induced the second extracellular signal‐regulated kinases 1 and 2 activation, resulting in posttranscriptional profibrotic gene expression. Conclusions These results demonstrate that extracellular signal‐regulated kinases 1 and 2 activation is important for Krüppel‐like factor 15–mediated IL‐11 expression in adventitial fibroblasts to promote adventitial remodeling in Ang II–induced hypertension. Therefore, targeting the Krüppel‐like factor 15/IL‐11 axis might serve as a new therapeutic strategy for vascular diseases.

Herbivore-induced plant volatiles (HIPVs) have been associated with plant-plant-herbivorous-natural enemies communication and an enhanced response to the subsequent attack. Spodoptera litura is a serious cosmopolitan pest that has developed a high level of resistance to many insecticides. However, the underlying molecular and biochemical mechanism by which HIPV priming reduces S. litura larval sensitivity to insecticides remains largely unknown. This study was conducted to explore the potential of volatile from undamaged, or artificially damaged, or S. litura-damaged tomato plants on the susceptibility of S. litura to the insecticides beta-cypermethrin indoxacarb and chlorpyrifos. We found that larvae exposed to volatile from S. litura-damaged or artificially damaged tomato plants were significantly less susceptible to the three insecticides than those exposed to volatile from undamaged tomato plants. Elevated activities of detoxifying enzymes [cytochrome P450 monooxygenases (P450s), glutathione S-transferases (GSTs), and esterases (ESTs)], were expressed in S. litura larvae exposed to volatile from S. litura-damaged tomato plants than those exposed to volatile from undamaged tomato plants. Similarly, seven detoxification-related genes [GSTs (SlGSTe1, SlGSTo1, and SlGSTe3) and P450s (CYP6B48, CYP9A40, CYP321A7, and CYP321B1)] in the midgut and fat body of larvae were up-regulated under exposure to volatile from S. litura-damaged tomato plants. Increased volatile organic compounds emissions were detected in the headspace of tomato plants damaged by S. litura compared to the undamaged plants. Collectively, these findings suggest that HIPVs can considerably reduce caterpillar susceptibility to insecticides, possibly through induction-enhanced detoxification mechanisms, and provide valuable information for implementing an effective integrated pest management strategy.

Nonalcoholic steatohepatitis (NASH) has been linked to inflammation induced by intestinal microbiota. Poria cocos polysaccharides (PCP) possesses anti-inflammation and immunomodulation functions; however, its preventive effects against NASH and potential mechanisms need to be explored.The composition of PCP was determined using ion chromatography. C57BL/6 mice were administered the methionine and choline deficient (MCD) diet for 4 weeks to establish the NASH model or methionine-choline-sufficient (MCS) diet to serve as the control. Mice were assigned to the MCS group, MCD group, low-dose PCP (LP) group, and high-dose PCP (HP) group, and were administered the corresponding medications via gavage. Serum biochemical index analysis and liver histopathology examination were performed to verify the successful establishment of NASH model and to evaluate the efficacy of PCP. The composition of intestinal bacteria was profiled through 16S rRNA gene sequencing. Hepatic RNA sequencing (RNA-Seq) was performed to explore the potential mechanisms, which were further confirmed using qPCR, western blot, and immunohistochemistry.PCP consists of glucose, galactose, mannose, D-glucosamine hydrochloride, xylose, arabinose, and fucose. PCP could significantly alleviate symptoms of NASH, including histological liver damage, impaired hepatic function, and increased oxidative stress. Meanwhile, HP could reshape the composition of intestinal bacteria by significantly increasing the relative abundance of Faecalibaculum and decreasing the level of endotoxin load derived from gut bacteria. PCP could also downregulate the expression of pathways associated with immunity and inflammation, including the chemokine signaling pathway, Toll-like receptor signaling pathway, and NF-kappa B signaling pathway. The expression levels of CCL3 and CCR1 (involved in the chemokine signaling pathway), Tlr4, Cd11b, and NF-κb (involved in the NF-kappa B signaling pathway), and Tnf-α (involved in the TNF signaling pathway) were significantly reduced in the HP group compared to the MCD group.PCP could prevent the development of NASH, which may be associated with the modulation of intestinal microbiota and the downregulation of the NF-κB/CCL3/CCR1 axis.

Context Laurolitsine is an aporphine alkaloid and exhibits potent antihyperglycemic and antihyperlipidemic effects in ob/ob mice. Objective To investigate the pharmacokinetics, tissue distribution and excretion of laurolitsine. Materials and methods A LC-MS/MS method was established and validated to determine laurolitsine concentrations in the biological matrix of rats (plasma, tissue homogenate, urine and faeces). 10 Sprague-Dawley (SD) rats were used for plasma exposure study: 5 rats were injected with 2.0 mg/kg of laurolitsine via the tail vein, and the other 5 rats were administered laurolitsine (10.0 mg/kg) by gavage. 25 SD rats used for tissue distribution study and 5 SD rats for urine and faeces excretion study: rats administered laurolitsine (10.0 mg/kg) by gavage. After administered, serial blood, tissue, urine and faeces were collected. Analytical quantification was performed by a previous LC-MS/MS method. The pharmacokinetics, bioavailability, tissue distribution and excretion of laurolitsine were described. Results The pharmacokinetic parameters of oral and intravenous administration with Tmaxwere 0.47 and 0.083 h, t1/2 were 3.73 and 1.67 h, respectively. Oral bioavailability was as low as 18.17%. Laurolitsine was found at a high concentration in the gastrointestinal tract, liver, lungs and kidneys (26 015.33, 905.12, 442.32 and 214.99 ng/g at 0.5 h, respectively) and low excretion to parent laurolitsine in urine and faeces (0.03 and 1.20% in 36 h, respectively). Conclusions This study established a simple, rapid and accurate LC-MS/MS method to determine laurolitsine in different rat samples and successful application in a pharmacokinetic study.

Rationale: Data on the molecular mechanisms that regulate platelet–pulmonary endothelial adhesion under conditions of hypoxia are lacking, but may have important therapeutic implications. Objectives: To identify a hypoxia-sensitive, modifiable mediator of platelet–pulmonary artery endothelial cell adhesion and thrombotic remodeling. Methods: Network medicine was used to profile protein–protein interactions in hypoxia-treated human pulmonary artery endothelial cells. Data from liquid chromatography–mass spectrometry and microscale thermophoresis informed the development of a novel antibody (Ab) to inhibit platelet–endothelial adhesion, which was tested in cells from patients with chronic thromboembolic pulmonary hypertension (CTEPH) and three animal models in vivo. Measurements and Main Results: The protein NEDD9 was identified in the hypoxia thrombosome network in silico. Compared with normoxia, hypoxia (0.2% O(2)) for 24 hours increased HIF-1α (hypoxia-inducible factor-1α)–dependent NEDD9 upregulation in vitro. Increased NEDD9 was localized to the plasma-membrane surface of cells from control donors and patients with CTEPH. In endarterectomy specimens, NEDD9 colocalized with the platelet surface adhesion molecule P-selectin. Our custom-made anti-NEDD9 Ab targeted the NEDD9–P-selectin interaction and inhibited the adhesion of activated platelets to pulmonary artery endothelial cells from control donors in vitro and from patients with CTEPH ex vivo. Compared with control mice, platelet–pulmonary endothelial aggregates and pulmonary hypertension induced by ADP were decreased in NEDD9(−/−) mice or wild-type mice treated with the anti-NEDD9 Ab, which also decreased chronic pulmonary thromboembolic remodeling in vivo. Conclusions: The NEDD9–P-selectin protein–protein interaction is a modifiable target with which to inhibit platelet–pulmonary endothelial adhesion and thromboembolic vascular remodeling, with potential therapeutic implications for patients with disorders of increased hypoxia signaling pathways, including CTEPH.

Purpose This study was designed to probe the effect of downhill running on microtubule acetylation and autophagic flux in rat skeletal muscle. Methods Sprague-Dawley rats were subjected to an exercise protocol of a 90-min downhill run with a slope of -16° and a speed of 16 m/min, and then the soleus was sampled at 0 h, 12 h, 24 h, 48 h, and 72 h after exercise. Protein expression levels of microtubule-associated protein 1 light chain 3 (LC3), p62/sequestosome 1 (p62), α-tubulin and acetylated α-tubulin (AcK40 α-tubulin) were detected by Western blotting. Alpha-tubulin was costained with acetylated α-tubulin (AcK40 α-tubulin) or cytoplasmic dynein intermediate chain in a single muscle fiber, and LC3 was costained with lysosomal associated membrane protein 1 (LAMP1) in cryosections. To assess autophagic flux in vivo, colchicine or vehicle was injected intraperitoneally 3 days before the exercise experiment, and the protein levels of LC3 and p62 were measured by Western blotting. Results Downhill running induced a significant increase in the protein levels of LC3-II and p62, while the level and proportion of AcK40 α-tubulin were markedly decreased. Furthermore, the amount of dynein on α-tubulin was decreased after downhill running, and autophagosomes accumulated in the middle of myofibrils. Importantly, LC3-II flux was decreased following downhill running compared with that in the control group. Conclusions A bout of downhill running decreases microtubule acetylation, which may impair dynein recruitment and autophagosome transportation, causing blocked autophagic flux.

Human serum albumin (HSA)-based therapeutics have attracted tremendous attention in the development of anticancer agents. The versatile properties of HSA make HSA-based therapeutics possess improved pharmacokinetics, extended circulation half-life, enhanced efficacy, reduced toxicity, etc. Generally, the HSA-based therapeutics systems can be divided into four categories, i.e. HSA-drug nanoparticles, HSA-drug conjugates, HSA-binding prodrugs, and HSA-based recombinant fusion proteins: the latter mainly include antibody (domain)- and cytokine- fusion proteins. Advances in this area revealed the advantages of HSA-based systems in the development of tumor site-oriented therapeutics, partly referring to the enhanced penetration and retention (EPR) effect and the intensive macropinocytosis. Accordingly, a variety of technical platforms for the design and preparation of HSA-based therapeutics have been reported. Major strategies and directions for the drug development were discussed; those include (1) Tumor-site oriented drug delivery and enhanced drug retention, (2) Tumor-site prodrug release and activation, (3) Cancer cell bound intensive drug internalization, and (4) Tumor microenvironment (TME) directed immunomodulation. Notably, the multimodal HSA-based approach is promising for the development of tumor-oriented therapeutics for cancer therapy.

To observe the changes of the skeletal muscle fibrosis and changes of transforming growth factor-β1(TGF-β1)/ extracellular signal-regulated kinase (ERK) / connective tissue growth factor (CTGF)pathway in rats after long-term eccentric exercise and acupuncture intervention, so as to explore the mechanism of acupuncture in regulating exercise-induced skeletal muscle fibrosis.A total of 30 male SD rats were randomly divided into normal control, exercise and acupuncture group, with 10 rats in each group. The rat model of skeletal muscle fibrosis was established by eccentric exercise for 3 weeks. After exercise trained every time, the rats of the acupuncture group received acupuncture stimulation by holding the acupuncture needle to obliquely and longitudinally penetrate the ventral part of triceps of the lower leg along its lateral side, followed by retaining the needle for 2 min. Changes of the collagen fibers in each group was observed by scanning electron microscope. The expressions of Collagen Ⅰ, TGF-β1, phosphated (p)-ERK/ERK and CTGF proteins were detected by Western blot.After 3 weeks of eccentric exercise and acupuncture, the fibrosis and deposition of collagen fibers in the exercise group were significantly higher than that in the normal control group,the degree of fibrosis in the acupuncture group was significantly lower than that in the exercise group. Compared with the normal control group, the expression levels of Collagen Ⅰ, TGF-β1, CTGF and p-ERK/ERK in the exercise group was significantly higher (The accumulation of chronic sports injury can lead to the deposition of collagen fibers in skeletal muscle, which leads to the fibrosis of skeletal muscle. Acupuncture can inhibit skeletal muscle fibrosis via down-regulating TGF-β1/ERK/CTGF signaling pathway.

Aims Glia-mediated neuro-inflammation and oxidative stress-induced neuronal apoptosis can contribute to epileptogenesis. We have demonstrated previously that mimetics of glucagon-like peptide-1 (GLP-1), glucose-dependent insulinotropic polypeptide (GIP) and dual-GLP-1/GIP receptor agonists protect the brain from inflammation, oxidative stress, apoptosis and neuronal loss in animal models of central nervous system diseases. Methods This study investigated for the first time whether the novel dual GLP-1/GIP receptor agonist DA3-CH has neuroprotective effects in the pilocarpine-induced status epilepticus (SE) rat model and the studies the underlying mechanisms. DA3-CH was administered once daily at 10 nmol/kg ip. following SE induction. The effect of DA3-CH was evaluated by immunohistochemistry and western blot at 12 h, 1 d, 3 d, 7 d after kindling. Results Our findings show that DA3-CH reduced the chronic inflammation response (astrogliosis and microgliosis), and the associated release of the pro-inflammatory cytokines interleukin-1β (IL-β) and tumor necrosis factor-α (TNF-α) in the hippocampal CA1 area. Furthermore, DA3-CH reduced the expression of the mitochondrial pro-apoptotic protein Bax, while increasing the expression of the anti-apoptotic protein Bcl-2. Neuronal numbers in the CA1 area were much reduced by pilocarpine treatment, and DA3-CH protected neurons from neurotoxicity. Conclusion These results demonstrated that DA3-CH could mitigate pilocarpine-induced neuro-inflammation, mitochondrial apoptosis and neuronal loss. The findings encourage the development of dual agonists as novel therapeutic interventions for epilepsy.

There is substantial evidence supporting the notion that the primary somatosensory (S1) cortex is an important structure involved in the perceptional component of pain. However, investigations have mainly focused on other pain-related formations, and few reports have been provided to investigate the synaptic plasticity in the S1 cortex in response to persistent pain. In the present study, we report that bee venom (BV) injection triggered an imbalance between excitatory and inhibitory synaptic transmission in the S1 cortex in rats. Using a multi-electrode array recording, we found that BV-induced persistent inflammatory pain led to temporal and spatial enhancement of synaptic plasticity. Moreover, slice patch clamp recordings on identified pyramidal neurons demonstrated that BV injection increased presynaptic and postsynaptic transmission in excitatory synapses and decreased postsynaptic transmission in inhibitory synapses in the layer II/III neurons within the S1 cortex. In immunohistochemistry and Western blot sections, the distribution and expression of total AMPA receptor subunits and gamma-amino butyric acid-A (GABAA) were unaffected, although the membrane fractions of GluR2 and GABAA were decreased, and their cytosolic fractions were increased in contrast. The change of GluR1 was opposite to that of GluR2, and GluR3 did not change significantly. Our studies, therefore, provide direct evidence for both presynaptic and postsynaptic changes in synapses within the S1 cortex in persistent nociception, which are probably related to the membrane trafficking of GluR1, GluR2, and GABAA. Perspective: Increased synaptic plasticity was detected in S1 after peripheral nociception, with enhanced excitatory and decreased inhibitory synaptic transmissions. Increased GluR1, and decreased GABAAα1 and GluR2 membrane trafficking were detected. Therefore, the disrupted excitatory/inhibitory balance in transmissions is involved in nociception processing, and S1 can be a potential antinociceptive site.

Background Bee pollen has been regarded as a complete nutritional human dietary supplement but its nutrient absorption and biological effects may be restricted by the complex pollen wall. The aim of this study was to explore the effects of ultrasonic and ball-milling treatment on the release of nutritional components and on in vitro and in vivo antioxidant effects of rose (Rosa rugosa) bee pollen. Results Bee pollen walls were broken to varying degrees, nutrients were released, and in vitro and in vivo antioxidant effects of bee pollen were improved. The scavenging effects of 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2'-azinobis(3-ethylbenzothiazolone-6-sulfonic acid) (ABTS) radicals, and oxygen radical absorbance capacity (ORAC) were improved. In aging mice, wall-breaking treatment led to better organ recovery, enhanced superoxide dismutase (SOD) and catalase (CAT) effects, and malondialdehyde (MDA) reduction. Eight compounds of rose bee pollen ethanol extract, including isorhamnetin 3-O-diglucoside and N', N″, N‴-dicaffeoyl p-coumaroyl spermidine were identified by ultra-performance liquid chromatography electrospray ionization quadrupole time of flight mass spectrometry (UPLC-ESI-QTOF-MS/MS) assay. Conclusion This study showed that ultrasonic treatment had greater wall-disruption effects of bee pollen on nutrient release and antioxidant effect promotion. In conclusion, rose bee pollen, with wall-breaking treatments, may have potential value as an ingredient in functional food processing. © 2019 Society of Chemical Industry.

The isolated perfused rat lung (IPL), coupled with high performance liquid chromatography\tandem mass spectrometry analysis (HPLC–ESI-MSn), has been developed as a tool for screening bioactive components in Glycyrrhiza uralensis Fisch. (GU). First, IPL was perfused with the water extract of GU (EGU), the bioactive components in the EGU would selectively combine to the receptors or channels of lung. By changing the pH of perfused solution, the combined components were eluated and then detected by HPLC-ESI-MSn. Four compounds were detected in the desorption eluate of IPL, among these compounds, liquiritin (1), ononin (2) and glycyrrhizic acid (4) were identified by comparing with the chromatography of the standards, while licorice-saponin G2 (3) were determined by analysis of the structure clearage characterization of mass spectrometry. Then, due to the lack of compound 3 sample, compounds 1, 2 and 4 with respective concentrations of 50 μM, 5 μM, 500 nM, 50 nM and 5 nM were applied to evaluate the protective effect of pulmonary epithelial cells (PEC, A549 cell) injury induced by lipopolysaccharide (LPS) for anti-inflammatory activity assessment. The results showed that except the 5 nM group of compound 1, 5 nM and 50 nM groups of compound 2, all other groups could remarkably inhibit the PEC injury (vs LPS group, 2-500 nM groups: p

Although not without controversy, as a general trend, the human sperm count is declining world-wide. One major reason for such a decline is an increase in the human life-span. According to the life history tradeoff theory, fecundity is inversely related to the lifespan; the longer the lifespan, the lower the fecundity. This is essential to the maintainance of diversity and balance of different species. Such a corrleation validated by experimental data that show that the extension of life in Caenorhabditis elegans, Drosophila and Rodents is associated with reduction in fecundity. The demographic data from a public data source, shows that the total fertility rate is positively correlated with the infant death rate, it is inversely correlated with the life expectancy. We postulate that the fall in spermatogenesis might be regulated by the neuroendocrine system that underlie human longevity.

Multiple sclerosis, inflammatory bowel disease and organ transplant rejection are related to Th17 cell development and inflammatory respond. RORγt, a specific transcription factor regulating Th17 cell differentiation, is a pivotal target for the treatment of diseases. However, the clinical application of RORγt inverse agonists reported so far has been hindered due to limited efficacy and toxic side effects. Plant-derived natural products with drug-like properties and safety are wide and valuable resources for candidate drug discovery. Herein, structure-based virtual screening was used to find out 2',4'-Dihydroxy-2,3-dimethoxychalcone (DDC), a chalcone derivative rich in plants and food, located in the binding pocket of RORγt and targeted to inhibit RORγt activity. DDC repressed murine Th17 differentiation and promoted Treg differentiation remarkably in a dose-dependent manner. In addition, DDC treatment improved experimental autoimmune encephalomyelitis recovery, ameliorated experimental colitis severity, and prevented graft rejection significantly. Mechanically, DDC indirectly stabilized Foxp3 expression by inhibiting RORγt activity and the expression of its target gene profile in vitro and in vivo, which realized its regulation of Th17/Treg balance. In conclusion, our study provides a scientific basis that DDC, as an inverse agonist of RORγt with simple structure, rich sources, low cost, high efficiency, and low toxicity, has great potential for the development of a novel effective immunomodulator for the treatment of Th17-mediated inflammatory diseases.

The role of metallothionein (MT)-like cadmium (Cd) binding protein (MTLCdBP) in protecting the testes against Cd toxicity was examined. In the acute Cd exposure treatment, cadmium chloride was intraperitoneally injected at 2 mg Cd/kg to Wistar male rats. In the chronic Cd toxicity treatments, 20 mg Cd/kg/d was orally administered for 5 d a week for 5, 10, and 15 wk. MT (-I,-II) and MTLCdBP were measured using ELISA and Cd-Hem methods, respectively. Testicular tissues were immunostained with antibodies of MT-I,-II, MT-III, and MTLCdBP. Expression of HO1, OGG, iNOS, COX2, and p53 was measured by RT-PCR. Cd concentration in the testis increased dose-dependently in response to Cd exposure. MTLCdBP concentration increased markedly with increasing Cd accumulation. Significant increases in expression of iNOS, HO1, COX2, and OGG1 were observed in the acute exposure treatment. In the chronic oral administration group, expression of MT-I, MT-II, MT-III, iNOS, HO1, and COX2 did not change. Positive immunostaining of MTLCdBP was observed in testicular interstitial tissue. In the testis protected from Cd toxicity, MTLCdBP induction increased significantly with increasing Cd accumulation. Our results suggest that MTLCdBP plays an important role in protecting the testis against Cd toxicity.

Talaromycosis (penicilliosis) caused by Talaromyces marneffei is one of the most important opportunistic infection diseases in tropical countries of South and Southeast Asia. Most infections occurred in individuals with human immunodeficiency virus (HIV) and the primarily reason for the increase in the number of the cases is HIV pandemic. The pathogenesis of T. marneffei infection is unclear. There is still no ideal animal model for studying talaromycosis. In this study, we developed a stable, safe and maneuverable murine model that mimics human T. marneffei disseminated infection using T. marneffei yeast intraperitoneal injected to BALB/c nude mice. We successfully observed symptoms similar to those seen in clinical patients in this murine model, including skin lesions, hepatosplenomegaly, pulmonary infection and mesenteric lesions. We further studied the pathological changes of various tissues and organs in the infected animals to help better understand the severity of the infection. This model may provide a good tool for studying disseminated infection induced by T. marneffei.

Background Long non-coding RNAs (lncRNAs) play vital roles in the development and progression of non-small-cell lung cancer (NSCLC); however, the role of most lncRNAs in NSCLC remains unknown. This study explored the clinical significance, biological function and underlying mechanism of lnc-GAN1 in NSCLC. Methods With a custom lncRNA microarray we found that lnc-GAN1 is markedly downregulated in NSCLC tissues. Then lnc-GAN1 expression level was measured using qRT-PCR in NSCLC tissues and cell lines. Survival was assessed using the Kaplan-Meier method. The biological functions of lnc-GAN1 in lung cancer cells were evaluated in vitro and in vivo. RNA fluorescence in situ hybridization and subcellular localization assays revealed the subcellular distribution of lnc-GAN1 in cells. Bioinformatic analysis was adopted to predict miRNAs and signaling pathways regulated by lnc-GAN1. RNA immunoprecipitation and Dual-luciferase reporter assays were used to assess the interaction between lnc-GAN1 and miR-26a-5p in lung cancer cells. Results lnc-GAN1 is downregulated in HCC tissues and associated with larger tumor size and poor overall survival and disease-free survival; its ectopic expression suppresses cell proliferation, colony formation, and cell cycle progression and induces apoptosis in NSCLC cells; it also inhibits tumor growth in the NSCLC xenograft model. We further proved that lnc-GAN1 is localized in cytoplasm and transcribed independently from its parental gene GAN. Mechanistically, lnc-GAN1 acts as a sponge for miR-26a-5p by two seed sequences, and the two non-coding RNAs have a negative relationship in NSCLC tissues; we further prove that PTEN is a direct target of miR-26a-5p and lnc-GAN1 inhibits cell cycle signaling pathway by activating PTEN, whose expression level correlated negatively with miR-26a-5p level but positively with lnc-GAN1 level in NSCLC samples. Conclusions Lnc-GAN1 is downregulated and associated with poor survival of NSCLC patients, and mechanistically acts as a tumor suppressor via sponging and inhibiting miR-26a-5p to upregulate PTEN. This study provides a potential prognostic biomarker and treatment target for NSCLC.

Recently, accumulating evidence revealed that nonalcoholic fatty liver disease (NAFLD) is highly associated with the dysbiosis of gut microbiota. Jiang Zhi Granule (JZG), which is composed of five widely used Chinese herbs, has shown hypolipidemic effect, while whether such effect is mediated by gut microbiota is still unclear. Here, we found that both low and high doses of JZG (LJZ and HJZ) could improve hepatic steatosis and function, as well as insulin resistance in NAFLD mice. 16S rRNA gene sequencing revealed that JZG treatment could reverse the dysbiosis of intestinal flora in NAFLD mice, exhibiting a dose-dependent effect. Notably, HJZ could significantly reduce the relative abundance of Desulfovibrionaceae, while increasing the relative abundance of such as S24_7 and Lachnospiraceae. PICRUSt analysis showed that HJZ could significantly alter the functional profile of gut microbiota, including the reduction of the lipopolysaccharide biosynthesis and sulfur metabolism pathway, which is verified by the decreased levels of fecal hydrogen sulfide (H2S) and serum lipopolysaccharide binding protein (LBP). In addition, hepatic mRNA sequencing further indicated that the HJZ group can regulate the peroxisome proliferator-activated receptor (PPAR) pathway and inflammatory signaling pathway, as validated by RT-PCR and Western blot. We also found that different doses of JZG may regulate lipid metabolism through differentiated pathways, as LJZ mainly through the promotion of hepatic lipid hydrolysis, while HJZ mainly through the improvement of hepatic lipid oxidation. Taken together, JZG could modulate gut dysbiosis with dose-effect, alleviate inflammation level, and regulate hepatic lipid metabolism, which may subsequently contribute to the improvement of NAFLD. Our study revealed the underlying mechanisms in the improvement of NAFLD by a Chinese herbal compound, providing future guidance for clinical usage.

To investigate the effects of high-load eccentric exercise on the ultrastructure of autophagy and the autophagy-related proteins Beclin1 and LC3II / I in rats.Forty-eight SD male rats were randomly divided into control group (C,The number of soleus muscle autophagosomes in group E was increased at 0, 12 and 24 hours after exercise, and LC3 autophagic fluorescence was significantly increased (High-load eccentric exercise can induce ultrastructural changes in skeletal muscle autophagy and increase the expression of autophagy protein. The peak value appears at 12 hours after exercise. The above may be one of the reasons for the decline in skeletal muscle function caused by sports injury.目的: 观察大负荷离心运动对大鼠骨骼肌自噬超微结构及自噬相关蛋白Beclin1和LC3II/I的影响。方法: 48只SD雄性大鼠适应性训练后随机分成对照组(C

Idiopathic pulmonary fibrosis (IPF) is a chronic, fatal lung disease characterized by progressive and non-reversible abnormal matrix deposition in lung parenchyma. Myofibroblasts originating mainly from resident fibroblasts via fibroblast-to-myofibroblast transition (FMT) are the dominant collagen-producing cells in pulmonary fibrosis. N(6)-methyladenosine (m(6)A) modification has been implicated in various biological processes. However, the role of m(6)A modification in pulmonary fibrosis remains elusive. In this study, we reveal that m(6)A modification is upregulated in a bleomycin (BLM)-induced pulmonary fibrosis mouse model, FMT-derived myofibroblasts, and IPF patient lung samples. Lowering m(6)A levels through silencing methyltransferase-like 3 (METTL3) inhibits the FMT process in vitro and in vivo. Mechanistically, KCNH6 is involved in the m(6)A-regulated FMT process. m(6)A modification regulates the expression of KCNH6 by modulating its translation in a YTH-domain family 1 (YTHDF1)-dependent manner. Together, our study highlights the critical role of m(6)A modification in pulmonary fibrosis. Manipulation of m(6)A modification through targeting METTL3 may become a promising strategy for the treatment of pulmonary fibrosis.

The aim of the present study was to investigate the effects of exercises with different durations and intensities on mitochondrial autophagy and FUNDC1 in rat skeletal muscles. Sixty male Sprague-Dawley rats were randomly divided into 2- and 4-week control groups (Con), moderate-intensity exercise groups (M-ex groups, treadmill exercise, 16 m/min, 1 h/d, 6 d/week), and high-intensity exercise groups (Hi-ex groups, treadmill exercise, 35 m/min, 20 min/d, 6 d/week). The bilateral soleus muscles were separated after the intervention, and paraffin sections were prepared for transmission electron microscopy. ELISA method was used to detect the content of citrate synthase (CS). The co-localizations of microtubule-associated protein 1 light chain 3 (LC3)/cytochrome c oxidase IV (COX-IV), FUNDC1/COX-IV and LC3/FUNDC1 were observed by immunofluorescent staining in frozen sections. The skeletal muscle mitochondria were extracted, and the expression of autophagy-related proteins, including AMPKα, p-AMPKα, Unc-51 like kinase 1 (ULK1), FUNDC1, LC3 and p62, were detected by Western blot. The results showed that exercise increased mitochondrial function, i.e. peroxisome proliferator-activated receptor γ co-activator-1α (PGC-1α), COX-I protein expression levels and CS content. There was no difference of mitochondrial function parameters between 2-week M-ex and 2-week Hi-ex groups, while mitochondrial function of 4-weeks Hi-ex group was significantly lower than that of 4-week M-ex group. Under the same exercise intensity, mitochondrial autophagy activation in skeletal muscle of 4-week exercise was higher than that in 2-week exercise group; Under the same duration of exercise, mitochondrial autophagy activation of Hi-ex group was higher than that in M-ex group. Both 2- and 4-week exercise intervention increased LC3/COX-IV, COX-IV/FUNDC1, and FUNDC1/LC3 co-localizations. Exercise increased LC3-II/LC3-I ratio, down-regulated p62 protein expression level, up-regulated FUNDC1, ULK1 protein expression levels and AMPKα phosphorylation, and the changes of these proteins in 4-week Hi-ex group were significantly greater than those in 4-week M-ex group. These results suggest exercise induces mitochondrial autophagy in skeletal muscles, and the activity of autophagy is related to the duration and intensity of exercise. The induction mechanism of exercise may involve the mediation of FUNDC1 expression through AMPK-ULK1 pathway.

To investigate the expression of apoptosis-related proteins Fas and FasL in the brain tissue of rats with traumatic brain injury and the effect of electroacupuncture on the expression of Fas and FasL, so as to explore the effective time window of electroacupuncture in the treatment of traumatic brain injury.Sprague-Dawley rats were randomly divided into blank group, sham-operation group, model group, and electroacupuncture treatment groups 1, 2, and 3. Traumatic brain injury was induced by the modified Feeney free-fall impact device, and for the rats in the electroacupuncture treatment groups 1, 2, and 3, electroacupuncture started at 4 hours and on days 3 and 7, respectively, after modeling and lasted to day 14. The Morris water maze test was used to evaluate learning and memory ability, and immunofluorescence assay and Western blot were used to observe the changes in the expression of Fas and FasL in traumatic brain tissue.Compared with the blank group and the sham-operation group, the model group had a lower percentage of time spent in the target quadrant from the 3rd day folowing modeling; after electroacupuncture intervention, the electroacupuncture treatment groups showed a gradual increase in the time spent in the target quadrant, and on day 7,10 and 14, electroacupuncture treatment group 1 had a significantly higher percentage than the model group (Early electroacupuncture intervention can regulate the apoptosis receptor pathway by down-regulating Fas and FasL to exert a therapeutic effect on traumatic brain injury and help with the recovery of cognition and memory ability after traumatic brain injury.

To observe the effect of electroacupuncture (EA) on neuronal apoptosis in rats with traumatic brain injury (TBI), and to explore the action mechanism of EA on improving the brain nerve function of TBI.A total of 88 6-week-old SD rats were randomly divided into a sham operation group, a model group, an EA group and a LY294002+EA group, 22 rats in each group. The TBI model on the left side was established by the improved Feeney's free fall method. After modeling for 24 h, the rats in the EA group and LY294002+EA group were treated with acupuncture at "Baihui" (GV 20) for 10 min and pricking acupuncture at "Shuigou" (GV 26) for 20 s; EA was applied at "Neiguan" (PC 6) and "Zusanli" (ST 36) on the right side (discontinuous wave, 2 Hz of frequency, 1 mA of intensity) for 10 min, once a day for 3 days. After 3 days of intervention, the TUNEL method was used to detect the level of neuron apoptosis in left cerebral cortex; the Western blot method was used to detect the expression of Akt, p-Akt, Bcl-2, Bax, Cyt-C and Caspase-9 in the left cerebral cortex.After 3-day treatment, compared with the sham group, the number of neuronal apoptosis in the left cortex was increased in the model group (EA could significantly reduce the neuronal apoptosis in rats with TBI, and its mechanism may be related to the activation of PI3K/Akt signaling pathway.

To observe the effect of electroacupuncture on the expression of phosphorylated adenosine 5'-monophosphate-activated protein kinase(p-AMPK), phosphorylated mammalian target of rapamycin(p-mTOR) and phosphorylated Ulk1(p-Ulk1) proteins in the cortex of traumatic brain injury (TBI) rats, so as to explore its mechanisms underlying treatment of TBI.Male SD rats were randomly divided into sham operation (sham), model, electroacupuncture Ⅰ (EA Ⅰ), electroacupuncture Ⅱ (EA Ⅱ) groups (After modeling and compared with the sham group, a large number of tissue necrosis, scattered arrangement of nerve fibers, vacuolar changes of cells, nuclear fragmentation, consolidation and hyperplastic scar tissue were found in the brain trauma area of rats in the model group. Nissl corpuscles were obviously absent. The ratio of p-AMPK/AMPK was up-regulated in the cortex of the wound region (Electroacupuncture may inhibit the over-activation of autophagy of cranial neurons by regulating the activation of AMPK, mTOR and Ulk1, thus exerting brain protection effect on TBI rats, and early electroacupuncture intervention is more effective in acute phase of TBI.

To observe the impacts of electroacupuncture (EA) on neurological function, the pathological morphology in brain tissue, apoptosis level and the protein expressions of apoptosis-related cytochrome C (Cyt-C) and cysteine aspartic acid protease-9 (Caspase-9) in the rats with traumatic brain injury (TBI) and explore the potential mechanism of EA in treatment of TBI.A total of 70 clean-grade SD mice were randomized into a blank group (8 rats), a sham-operation group (8 rats), a model group (27 rats) and an EA group (27 rats). In terms of interventions of 3, 7 and 14 days, 3 subgroups were divided in the model group and the EA group successively, 9 rats in each subgroup. The modified Feeney free-fall percussion method was adopted to establish TBI models of rats. In the sham-operation group, only the skull was exposed and drilled and no free-fall percussion was exerted. One day after modeling, EA was given in the rats of EA group at "Shuigou" (GV 26), "Baihui" (GV 20) and "Neiguan" (PC 6) and "Zusanli" (ST 36) on the affected side, with intermittent wave, 2 Hz in frequency, once daily, 10 min each time, for 3, 7 and 14 days successively. Separately, on the day 3, 7 and 14 of intervention, the modified neurological severity scale (mNSS) was used to evaluate the degree of neurological function injury in the rats, HE staining and Nissl staining were to observe the pathological and morphological changes in brain tissue, TUNEL method was to observe the level of apoptosis in brain tissue and immunohistochemistry (IHC) method and Western blot were to determine the protein expressions of Cyt-C and Caspase-9 in brain tissue.Compared with the sham-operation group, on the day 3, 7 and 14 of intervention, mNSS scores were increased obviously in the rats of the model group respectively (Electroacupuncture remarkably improves the condition in the neurological function injury and reduces apoptosis degree in TBI model rats, which is likely related to the down-regulation of the protein expressions of Cyt-C and Caspase-9 in damaged brain tissue and further to bring the impacts on mitochondria mediated apoptosis process.

We have shown previously that diallyl trisulfide (DATS) ameliorates mitochondrial fission and oxidative stress in a hyperglycemia-induced endothelial apoptosis and diabetic mouse model. The aim of this study was to investigate whether DATS mitigates Ang II-induced vascular smooth muscle cell (VSMC) phenotypic switching and vascular remodeling, and if so, to determine the underlying molecular events. Male C57BL/6 mice were used to establish a vascular remodeling model by continuous 2-week Ang II infusion using a subcutaneous osmotic pump. Animals were intraperitoneally injected with DATS or vehicle. Physiological parameters, vascular morphology, and molecular markers were assessed. For in vitro studies, VSMCs were pretreated with or without DATS for 1 h, then were stimulated with Ang II, and mitochondrial morphology and phenotypic switching of VSMCs were also measured. In primary mouse VSMCs, we found that Drp1-dependent mitochondrial fission regulated mitochondrial reactive oxygen species (mtROS) generation, which eventually promoted Ang II-induced VSMC proliferation, migration, and phenotypic switching. Moreover, Ang II was found to up-regulate the Rho-associated coiled coil-containing protein kinase 1 (ROCK1), which regulated mitochondrial fission and VSMC phenotypic switching by phosphorylating Drp1. However, the biological effect of Ang II was abrogated by DATS. Consistent with the effects in VSMCs, we found that DATS markedly alleviated mitochondrial fission, VSMC differentiation, and vessel wall thickening in an animal model of Ang II-induced vascular remodeling, which was regulated by the ROCK1/Drp1 signal. Our findings showed that DATS mitigated Ang II-induced vascular remodeling by suppressing Drp1-mediated mitochondrial fission in an ROCK1-dependent manner.