ABSTRACT Avian influenza A viruses generally do not replicate efficiently in human cells, but substitution of glutamic acid (Glu, E) for lysine (Lys, K) at residue 627 of avian influenza virus polymerase basic protein 2 (PB2) can serve to overcome host restriction and facilitate human infectivity. Although PB2 residue 627 is regarded as a species-specific signature of influenza A viruses, host restriction factors associated with PB2 627 E have yet to be fully investigated. We conducted immunoprecipitation, followed by differential proteomic analysis, to identify proteins associating with PB2 627 K (human signature) and PB2 627 E (avian signature) of influenza A/WSN/1933(H1N1) virus, and the results indicated that Tu elongation factor, mitochondrial (TUFM), had a higher binding affinity for PB2 627 E than PB2 627 K in transfected human cells. Stronger binding of TUFM to avian-signature PB2 590 G/ 591 Q and PB2 627 E in the 2009 swine-origin pandemic H1N1 and 2013 avian-origin H7N9 influenza A viruses was similarly observed. Viruses carrying avian-signature PB2 627 E demonstrated increased replication in TUFM-deficient cells, but viral replication decreased in cells overexpressing TUFM. Interestingly, the presence of TUFM specifically inhibited the replication of PB2 627 E viruses, but not PB2 627 K viruses. In addition, enhanced levels of interaction between TUFM and PB2 627 E were noted in the mitochondrial fraction of infected cells. Furthermore, TUFM-dependent autophagy was reduced in TUFM-deficient cells infected with PB2 627 E virus; however, autophagy remained consistent in PB2 627 K virus-infected cells. The results suggest that TUFM acts as a host restriction factor that impedes avian-signature influenza A virus replication in human cells in a manner that correlates with autophagy. IMPORTANCE An understanding of the mechanisms that influenza A viruses utilize to shift host tropism and the identification of host restriction factors that can limit infection are both critical to the prevention and control of emerging viruses that cross species barriers to target new hosts. Using a proteomic approach, we revealed a novel role for TUFM as a host restriction factor that exerts an inhibitory effect on avian-signature PB2 627 E influenza virus propagation in human cells. We further found that increased TUFM-dependent autophagy correlates with the inhibitory effect on avian-signature influenza virus replication and may serve as a key intrinsic mechanism to restrict avian influenza virus infection in humans. These findings provide new insight regarding the TUFM mitochondrial protein and may have important implications for the development of novel antiviral strategies.
Abstract Perfluorodecanoic acid (PFDoA) is a widely distributed environmental pollutant that can affect the functions of many organs. However, systematic evaluations of the effects of PFDoA on testicular functions are lacking. The aim of this study was to investigate the effects of PFDoA on mouse testicular functions, including spermatogenesis, testosterone synthesis, and stem Leydig cells (SLCs) in the interstitial tissue of the testis. PFDoA (0, 2, 5, 10 mg/kg/d) was administered via gavage to 2-month-old mice for 4 weeks. Serum hormone levels and sperm quality were assayed. Furthermore, to investigate the mechanisms by which PFDoA affects testosterone synthesis and spermatogenesis in vivo, the expression of StAR and P450scc in testicular tissue was measured by immunofluorescence staining and quantitative real-time PCR. In addition, the levels of SLC markers, including nestin and CD51, were studied. PFDoA decreased the luteinizing hormone concentration and sperm quality. Although the difference was not statistically significant, mean testosterone levels showed a downward trend. The expression of StAR, P450scc, CD51, and nestin was also suppressed in the PFDoA-treated groups compared with the control group. Our study suggested that PFDoA exposure can decrease testosterone biosynthesis, and even reduce the number of SLCs. These results indicated that PFDoA suppressed the main functions of testis, and further researches are required to identify strategies for preventing or reducing the effect of PFDoA on testicular function.
To study the chemical constituents from ethyl acetate extract of Micromelum falcatum.The constituents were separated and purified by silica gel, Sephadex LH-20 and HPLC. Their structures were elucidated by spectral analysis (NMR, MS).Ten compounds were isolated and identified as micropubescin (1), phebalosin (2), scopoletin (3), citrubuntin (4), thamnosmonin (5), hopeyhopin (6), arnottinin (7), casegravol (8), 2-methoxy-5-hydroxy cinnamate (9), threo-syringoylglycerol (10).Compounds 1 - 10 are obtained from this plant for the first time.
Objective To study the technological parameters of the purification process of phenols from Callicarpa nudiflora (C.nudiflora) with HP-20 macroporous resin.Methods The kinetic adsorption and desorption experiments were carried out on HP-20 macroporous resin (HP-20) to optimize the separation process of phenols.Additionally, the effects of four parameters including adsorption flow rate, elute flow rate, volume of 10% ethanol and 50% ethanol solution for elution were explored by a L4/3 orthogonal experiment.Finally, content of phenols in samples before and after being treated by HP-20 were compared.Results The results showed that a good separation and purification of crude total phenols extract from C. nudiflora was achieved using HP-20.The optimum conditions were found to be: initial concentration of 50 mg•mL -1 , absorb flow rate of 1 mL•min -1 , elute flow rate of 3 mL•min -1 , 5 BV of 10% ethanol solution for prewash and 10 BV of 50% ethanol solution for desorption solvent.Conclusion The content of phenols is above 40% in C. nudiflora after being treated by HP-20, indicating that HP-20 could be successfully applied to enrich and purify phenols in C. nudiflora.
Introduction: Zhixue Zhentong capsules (ZXZTCs) are a Tibetan medicine preparation solely composed of Lamiophlomis rotata (Benth.) Kudo. L. rotata is the only species of the genus Laniophlomis (family Lamiaceae) that has medicinal constituents derived from the grass or root and rhizome. L. rotata is one of the most extensively used folk medicines by Tibetan, Mongolian, Naxi, and other ethnic groups in China and has been listed as a first-class endangered Tibetan medicine. The biological effects of the plant include hemostasis, analgesia, and the removal of blood stasis and swelling. Purpose: This study aimed to profile the overall metabolites of ZXZTCs and those entering the blood. Moreover, the contents of six metabolites were measured and the hemostatic, analgesic, and anti-inflammatory effects of ZXZTCs were explored. Methods: Ultra-performance liquid chromatography-tandem quadrupole time-of-flight high-resolution mass spectrometry (UPLC-Q-TOF-MS) was employed for qualitative analysis of the metabolites of ZXZTCs and those entering the blood. Six metabolites of ZXZTCs were quantitatively determined via high-performance liquid chromatography The hemostatic, analgesic, and anti-inflammatory effects of ZXZTCs were evaluated in various animal models. Results: A total of 36 metabolites of ZXZTCs were identified, including 13 iridoid glycosides, 9 flavonoids, 9 phenylethanol glycosides, 4 phenylpropanoids, and 1 other metabolite. Overall, 11 metabolites of ZXZTCs entered the blood of normal rats. Quantitative analysis of the six main metabolites, shanzhiside methyl ester, chlorogenic acid, 8-O-acetyl shanzhiside methyl ester, forsythin B, luteoloside, and verbascoside, was extensively performed. ZXZTCs exerted hemostatic effects by reducing platelet aggregation and thrombosis and shortening bleeding time. Additionally, ZXZTCs clearly had an analgesic effect, as observed through the prolongation of the latency of writhing, reduction in writhing, and increase in the pain threshold of experimental rats. Furthermore, significant anti-inflammatory effects of ZXZTCs were observed, including a reduction in capillary permeability, the inhibition of foot swelling, and a reduction in the proliferation of granulation tissue. Conclusion: Speculative identification of the overall metabolites of ZXZTCs and those entering the blood can provide a foundation for determining its biologically active constituents. The established method is simple and reproducible and can help improve the quality control level of ZXZTCs as a medicinal product. Evaluating the hemostatic, analgesic, and anti-inflammatory activities of ZXZTCs can help reveal its mechanism.
Primary premature ejaculation (PPE) is a common male neurobiological disorder. Currently, there is consensus that the impairment in central serotonin (5-HT) neurotransmission constitutes a key pathogenic factor in PPE. Selective serotonin reuptake inhibitors (SSRIs) serve as the primary pharmacological intervention; however, a comprehensive elucidation of their mechanism of action remains incomplete. Owing to significant individual variability in efficacy, SSRIs exhibit a high discontinuation rate. Hence, there is an urgent need to address the selection of SSRIs for PPE treatment. This study aims to investigate the characteristics of tryptophan (TRP) metabolism in patients with PPE and to assess its influence on the efficacy of SSRIs. The exploratory study included a total of 16 patients with PPE and 16 control subjects who were healthy men without any sexual dysfunction. Upon enrollment in the study, all participants underwent a thorough medical history review and physical examination. Subsequently, their serum levels of TRP, its metabolites, large neutral amino acids (LNAAs), and metabolite ratios were assessed using a liquid chromatography-mass spectrometry (LC-MS) assay. After a period of 4 weeks of dapoxetine treatment, all patients with PPE underwent reassessment using the Premature Ejaculation Diagnostic Tool (PEDT) score and intravaginal ejaculatory latency time (IELT) test. The ratio of serum TRP to other LNAAs (TRP/LNAAs) in patients with PPE was found to be significantly lower compared to the control group (P < 0.05). Conversely, the ratio of kynurenine to TRP (KYN/TRP) was observed to be significantly higher in the PPE patients compared to the control group (P < 0.05). Including the serum TRP/LNAAs ratio and KYN/TRP ratio in the prediction model yielded the highest prediction efficiency for PPE. There was a significant negative correlation between the ratio of TRP/LNAAs before the treatment and the IELT after 4 weeks of the treatment. Additionally, there was a significant positive correlation observed between the ratio of TRP/LNAAs before the treatment and the PEDT score after 4 weeks of the treatment. This study demonstrates that the reduction in the TRP/LNAAs ratio and the elevation of the KYN/TRP ratio are significant characteristics associated with PPE. These findings suggest that diminished tryptophan availability in the brain and the activation of the kynurenine (KYN) pathway may play a role in the pathogenesis of PPE. The TRP/LNAAs ratio has potential as a reliable indicator of central serotonin (5-HT) levels. Considering the TRP/LNAAs ratio when selecting SSRIs for the treatment of PPE may enhance the response rate of this medication.
Over 70% land plants live in mutualistic symbiosis with arbuscular mycorrhizal (AM) fungi, and maintenance of symbiosis requires transcriptional and post-transcriptional regulation. The former has been widely studied, whereas the latter mediated by symbiotic microRNAs (miRNAs) remains obscure, especially in woody plants. Here, we performed high-throughput sequencing of the perennial woody citrus plant Poncirus trifoliata and identified 3750 differentially expressed genes (DEGs) and 42 miRNAs (DEmiRs) upon AM fungal colonization. By analyzing cis-regulatory elements in the promoters of the DEGs, we predicted 329 key AM transcription factors (TFs). A miRNA-mRNA regulatory network was then constructed by integrating these data. Several candidate miRNA families of P. trifoliata were identified whose members target known symbiotic genes, such as miR167h-AMT2;3 and miR156e-EXO70I, or key TFs, such as miR164d-NAC and miR477a-GRAS, thus are involved in AM symbiotic processes of fungal colonization, arbuscule development, nutrient exchange and phytohormone signaling. Finally, analysis of selected miRNA family revealed that a miR159b conserved in mycorrhizal plant species and a Poncirus-specific miR477a regulate AM symbiosis. The role of miR477a was likely to target GRAS family gene RAD1 in citrus plants. Our results not only revealed that miRNA-mRNA network analysis, especially miRNA-TF analysis, is effective in identifying miRNA family regulating AM symbiosis, but also shed light on miRNA-mediated post-transcriptional regulation of AM symbiosis in woody citrus plants.
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