Antibacterial activity of a polysaccharide isolated from Artemisia argyi leaf against Staphylococcus aureus and mechanism investigation.

Abuse of antibiotics has led to excessive amounts of antibiotic residues in food and environment, thus enhancing pathogenic bacterium resistance and threatening human health. Therefore, searching and developing safe and green antibiotic alternatives are necessary. In this study, an Artemisia argyi leaf polysaccharide (AALP) fraction was extracted and analyzed. Chemical composition analysis showed that the carbohydrate, uronic acid, protein, and polyphenol content in AALP were 68.3 % ± 4.13 %, 9.4 % ± 0.86 %, 1.79 % ± 0.27 %, and 0.16 % ± 0.035 %, respectively. Chromatographic results suggested that AALP contained rhamnose, arabinose, glucosamine, galactose, glucose, xylose, mannose, galacturonic acid, and glucuronic acid in a molar ratio of 9.26, 1.35, 1.18, 3.04, 48.51, 2.33, 31.26, 3.93, and 9.08; the weight average molecular weight, number average molecular weight, and polydispersity of AALP were 5.41 kDa, 4.63 kDa, and 1.168, respectively. Fourier transform infrared spectroscopy indicated that AALP constituted the polysaccharide-specific groups of CH, CO, and OH. Meanwhile, AALP showed a dose-dependent inhibitory effect on Staphylococcus aureus in the inhibition zone assay, and the minimal inhibitory concentration was 1.25 mg/mL. Furthermore, AALP disrupted the cell wall, depolarized the inner membrane potential, and inhibited the activities of succinate dehydrogenase and malate dehydrogenase in S. aureus.

A high-throughput screening assay for silencing established HIV-1 macrophage infection identifies nucleoside analogs that perturb H3K9me3 on proviral genomes.

HIV-infected macrophages are long-lived cells that represent a barrier to functional cure. Additionally, low-level viral expression by central nervous system (CNS) macrophages contributes to neurocognitive deficits that develop despite antiretroviral therapy (ART). We recently identified H3K9me3 as an atypical epigenetic mark associated with chronic HIV infection in macrophages. Thus, strategies are needed to suppress HIV-1 expression in macrophages, but the unique myeloid environment and the responsible macrophage/CNS-tropic strains require cell/strain-specific approaches. Here, we generated an HIV-1 reporter virus from a CNS-derived strain with intact auxiliary genes expressing destabilized luciferase. We employed this reporter virus in polyclonal infection of primary human monocyte-derived macrophages (MDM) for a high-throughput screen (HTS) to identify compounds that suppress virus expression from established macrophage infection. Screening ~6,000 known drugs and compounds yielded 214 hits. A secondary screen with 10-dose titration identified 24 meeting criteria for HIV-selective activity. Using three replication-competent CNS-derived macrophage-tropic HIV-1 isolates and viral gene expression readout in MDM, we confirmed the effect of three purine analogs, nelarabine, fludarabine, and entecavir, showing the suppression of HIV-1 expression from established macrophage infection. Nelarabine inhibited the formation of H3K9me3 on HIV genomes in macrophages. Thus, this novel HTS assay can identify suppressors of HIV-1 transcription in established macrophage infection, such as nucleoside analogs and HDAC inhibitors, which may be linked to H3K9me3 modification. This screen may be useful to identify new metabolic and epigenetic agents that ameliorate HIV-driven neuroinflammation in people on ART or prevent viral recrudescence from macrophage reservoirs in strategies to achieve ART-free remission. IMPORTANCE Macrophages infected by HIV-1 are a long-lived reservoir and a barrier in current efforts to achieve HIV cure and also contribute to neurocognitive complications in people despite antiretroviral therapy (ART). Silencing HIV expression in these cells would be of great value, but the regulation of HIV-1 in macrophages differs from T cells. We developed a novel high-throughput screen for compounds that can silence established infection of primary macrophages, and identified agents that downregulate virus expression and alter provirus epigenetic profiles. The significance of this assay is the potential to identify new drugs that act in the unique macrophage environment on relevant viral strains, which may contribute to adjunctive treatment for HIV-associated neurocognitive disorders and/or prevent viral rebound in efforts to achieve ART-free remission or cure.

Biocontrol potential and action mechanism of Bacillus amyloliquefaciens DB2 on Bipolaris sorokiniana.

Introduction: Bipolaris sorokiniana is the popular pathogenic fungi fungus which lead to common root rot and leaf spot on wheat. Generally, chemical fungicides are used to control diseases. However, the environmental pollution resulting from fungicides should not be ignored. It is important to study the mode of antagonistic action between biocontrol microbes and plant pathogens to design efficient biocontrol strategies. Results: An antagonistic bacterium DB2 was isolated and identified as Bacillus amyloliquefaciens. The inhibition rate of cell-free culture filtrate (CF, 20%, v/v) of DB2 against B. sorokiniana reached 92.67%. Light microscopy and scanning electron microscopy (SEM) showed that the CF significantly altered the mycelial morphology of B. sorokiniana and disrupted cellular integrity. Fluorescence microscopy showed that culture filtrate destroyed mycelial cell membrane integrity, decreased the mitochondrial transmembrane potential, induced reactive oxygen species (ROS) accumulation, and nuclear damage which caused cell death in B. sorokiniana. Moreover, the strain exhibited considerable production of protease and amylase, and showed a significant siderophore and indole-3-acetic acid (IAA) production. In the detached leaves and potted plants control assay, B. amyloliquefacien DB2 had remarkable inhibition activity against B. sorokiniana and the pot control efficacy was 75.22%. Furthermore, DB2 suspension had a significant promotion for wheat seedlings growth. Conclusion: B. amyloliquefaciens DB2 can be taken as a potential biocontrol agent to inhibit B. sorokiniana on wheat and promote wheat growth.

Fatty acid modification of antimicrobial peptide CGA-N9 and the combats against Candida albicans infection.

High-efficiency and low-toxic antimicrobial peptides (AMPs) are supposed to be the future candidates to solve the increasingly prominent problems of Candida albicans infection and drug resistance. Generally, introduction of hydrophobic moieties on AMPs resulted in analogues with remarkably increased activity against pathogens. CGA-N9, an antifungal peptide found in our lab, is a Candida-selective antimicrobial peptide capable of preferentially killing Candida spp. relative to benign microorganisms with low toxicities. We speculate that fatty acid modification could improve the anti-Candida activity of CGA-N9. In the present investigation, a set of CGA-N9 analogues with fatty acid conjugations at N-terminus were obtained. The biological activities of CGA-N9 analogues were determined. The results showed that the n-octanoic acid conjugation of CGA-N9 (CGA-N9-C8) was the optimal CGA-N9 analogue with the highest anti-Candida activity and biosafety; exhibited the strongest biofilm inhibition activity and biofilm eradication ability; and the highest stability against protease hydrolysis in serum. Furthermore, CGA-N9-C8 is less prone to develop resistance for C. albicans in reference with fluconazole; CGA-N9-C8 also exhibited Candidacidal activity to the planktonic cells and the persister cells of C. albicans; reduced C. albicans susceptibility in a systemic candidiasis mouse model. In conclusion, fatty acid modification is an effective method to enhance the antimicrobial activity of CGA-N9, and CGA-N9-C8 is a promising candidate to defend C. albicans infection and resolve C. albicans drug resistance.

Antifungal Activity and Plant Growth-Promoting Properties of Bacillus mojovensis B1302 against Rhizoctonia Cerealis.

Rhizoctonia cerealis is a worldwide soil-borne pathogenic fungus that significantly infects wheat and causes sharp eyespot in China. However, the biocontrol strains used for the control of Rhizoctonia cerealis are insufficient. In the present study, antagonistic strain B1302 from the rhizosphere of wheat were isolated and identified as Bacillus mojovensis based on their morphological, physiological, and biochemical characteristics, and their 16S rDNA sequence. Culture filtrate of strain B1302 had a broad antifungal spectrum. In order to improve the antifungal activity of B1302, response surface methodology (RSM) was used to optimize the culture conditions. The final medium composition and culture conditions were 13.2 g/L of wheat bran, 14.1 g/L of soybean meal, 224 r/min of rotation speed, 7.50 of initial pH, and 1.5 × 108 CFU/mL of inoculation amount at 35 °C for a culture duration of 72 h. B. mojavensis B1302 inhibited the hyphae growth of R.cerealis and produced hydrolytic enzymes (protease, chitinase, and glucanase), IAA, and had N-fixing potentiality and P-solubilisation capacity. It can also promote wheat seedling growth in potted plants. The disease incidence and index of wheat seedlings were consistent with the effect of commercial pesticides under treatment with culture filtrate. The biocontrol efficacy of culture filtrate was significant-up to 65.25%. An animal toxicological safety analysis suggested that culture filtrate was safe for use and could be developed into an effective microbial fungicide to control wheat sharp eyespot.

Antagonistic Activity and Mechanism of Bacillus subtilis XZ16-1 Suppression of Wheat Powdery Mildew and Growth Promotion of Wheat.

Wheat powdery mildew caused by Blumeria graminis f. sp. tritici (Bgt) is one of the most serious wheat diseases in the world. Biological control is considered an environmentally safe approach to control plant diseases. Here, to develop effective biocontrol agents for controlling wheat powdery mildew, antagonistic strain XZ16-1 was isolated and identified as Bacillus subtilis based on the morphological, biochemical, and physiological characteristics and 16S rDNA sequence. The culture filtrate of B. subtilis XZ16-1 and its extracts had a significant inhibitory effect on the spore germination of Bgt. Moreover, the therapeutic and prevention efficacy of the 100% culture filtrate on wheat powdery mildew reached 81.18 and 83.72%, respectively, which was better than that of chemical fungicide triadimefon. Further antimicrobial mechanism analysis showed that the XZ16-1 culture filtrate could inhibit the development of powdery mildew spores by disrupting the cell membrane integrity, causing reductions in the mitochondrial membrane potential, and inducing the accumulation of reactive oxygen species in the spores. Biochemical detection indicated that XZ16-1 could solubilize phosphate, fix nitrogen, and produce hydrolases, lipopeptides, siderophores, and indole-3-acetic acid. Defense-related enzymes activated in wheat seedlings treated with the culture filtrate indicated that disease resistance was induced in wheat to resist pathogens. Furthermore, a 106 CFU/ml suspension of XZ16-1 increased the height, root length, fresh weight, and dry weight of wheat seedlings by 77.13, 63.46, 76.73, and 19.16%, respectively, and showed good growth-promotion properties. This study investigates the antagonistic activity and reveals the action mechanism of XZ16-1, which can provide an effective microbial agent for controlling wheat powdery mildew.

Preparation of high-value porous carbon by microwave treatment of chili straw pyrolysis residue.

Microwave technology is utilized to prepare porous carbon from the chili straw pyrolysis residue in this study. As the pyrolysis temperature increases, the thermal stability of biochar is higher. The carbon speciation of the porous carbon PC500 is closest to that of graphite, and its inorganic-C reaches to 51.21%. Notably, the specific surface area of the activated porous carbon increases with increasing pyrolysis temperature, with a maximum value of 2768.52 m2/g for PC500. Further testing of the electrochemical properties of the porous carbon, PC500 possesses a high specific capacitance of 352F/g at 1 A/g while that of conventional heating is only 226.1F/g. The porous carbon prepared by microwave heating has better electrical properties compared to conventional heating, and the biochar obtained at higher pyrolysis temperature has a richer pore structure after activation.

Highly restricted SARS-CoV-2 receptor expression and resistance to infection by primary human monocytes and monocyte-derived macrophages.

Severe acute respiratory syndrome (SARS) coronavirus 2 (SARS-CoV2), which causes the disease COVID-19, has caused an unprecedented global pandemic. Angiotensin-converting enzyme 2 (ACE2) is the major cellular receptor for SARS-CoV2 entry, which is facilitated by viral Spike priming by cellular TMPRSS2. Macrophages play an important role in innate viral defense and are also involved in aberrant immune activation that occurs in COVID-19, and thus direct macrophage infection might contribute to severity of SARS-CoV2 infection. Here, we demonstrate that monocytes and monocyte-derived macrophages (MDM) under in vitro conditions express low-to-undetectable levels of ACE2 and TMPRSS2 and minimal coexpression. Expression of these receptors remained low in MDM induced to different subtypes such as unpolarized, M1 and M2 polarized. Untreated, unpolarized, M1 polarized, and M2 polarized MDM were all resistant to infection with SARS-CoV2 pseudotyped virions. These findings suggest that direct infection of myeloid cells is unlikely to be a major mechanism of SARS-CoV2 pathogenesis. Summary sentence: Monocytes and macrophages express minimal ACE2 and TMPRSS2 and resist SARS-CoV-2 Spike-mediated infection, suggesting direct myeloid cell infection is unlikely a major contributor to pathogenesis.

Epigenetic Landscape of HIV-1 Infection in Primary Human Macrophage.

Human immunodeficiency virus (HIV)-infected macrophages are long-lived cells that sustain persistent virus expression, which is both a barrier to viral eradication and contributor to neurological complications in patients despite antiretroviral therapy (ART). To better understand the regulation of HIV-1 in macrophages, we compared HIV-infected primary human monocyte-derived macrophages (MDM) to acutely infected primary CD4 T cells and Jurkat cells latently infected with HIV (JLAT 8.4). HIV genomes in MDM were actively transcribed despite enrichment with heterochromatin-associated H3K9me3 across the complete HIV genome in combination with elevated activation marks of H3K9ac and H3K27ac at the long terminal repeat (LTR). Macrophage patterns contrasted with JLAT cells, which showed conventional bivalent H3K4me3/H3K27me3, and acutely infected CD4 T cells, which showed an intermediate epigenotype. 5'-Methylcytosine (5mC) was enriched across the HIV genome in latently infected JLAT cells, while 5'-hydroxymethylcytosine (5hmC) was enriched in CD4 cells and MDMs. HIV infection induced multinucleation of MDMs along with DNA damage-associated p53 phosphorylation, as well as loss of TET2 and the nuclear redistribution of 5-hydoxymethylation. Taken together, our findings suggest that HIV induces a unique macrophage nuclear and transcriptional profile, and viral genomes are maintained in a noncanonical bivalent epigenetic state. IMPORTANCE Macrophages serve as a reservoir for long-term persistence and chronic production of HIV. We found an atypical epigenetic control of HIV in macrophages marked by heterochromatic H3K9me3 despite active viral transcription. HIV infection induced changes in macrophage nuclear morphology and epigenetic regulatory factors. These findings may identify new mechanisms to control chronic HIV expression in infected macrophages.

Study on sampling scheme for detecting mycotoxin during wheat storage.

BACKGROUND: Mycotoxin produced by mould is one of the most serious contamination sources in food security. Safe storage of grain has become more important to control food security. Currently, there is no officially approved or standardized sampling scheme for detecting mycotoxin in grain storage worldwide. RESULTS: In this study, deoxynivalenol (DON) was taken as a typical mycotoxin in stored wheat to be detected. Population density of corn weevil could not significantly increase wheat moisture, but wheat moisture was highly significantly and positively correlated with DON content (P < 0.01). Corn weevil density significantly increased the DON content in wheat. DON contamination degree was mainly distributed in the region of 14-20 cm below the surface layer of wheat. In the process of ventilation and dehumidification during the storage period, moisture of wheat decreased slightly with the extension of ventilation, but the DON content in wheat increased significantly. Combined with the analysis of ventilation, DON content in the upper layer and H1 position, where the wind direction is not easy to reach, increased significantly. CONCLUSION: Areas with high insect population density (14-20 cm below the surface layer of stored wheat) and low ventilation and high humidification (H1 position in the upper layer) should be taken as the key cutting sample areas for detecting mycotoxin during the period of grain storage. This study provides for the first time a scientific basis for the standardization of the wheat sampling scheme to monitor mycotoxin contamination during wheat storage. © 2022 Society of Chemical Industry.

Antagonistic Strain Bacillus amyloliquefaciens XZ34-1 for Controlling Bipolaris sorokiniana and Promoting Growth in Wheat.

Common root rot, caused by Bipolaris sorokiniana, is one of the most prevalent diseases of wheat and has led to major declines in wheat yield and quality worldwide. Here, strain XZ34-1 was isolated from soil and identified as Bacillus amyloliquefaciens based on the morphological, physiological, biochemical characteristics and 16S rDNA sequence. Culture filtrate (CF) of strain XZ34-1 showed a high inhibition rate against B.sorokiniana and had a broad antifungal spectrum. It also remarkably inhibited the mycelial growth and spore germination of B. sorokiniana. In pot control experiments, the incidence and disease index of common root rot in wheat seedlings were decreased after treatment with CF, and the biological control efficacy was significant, up to 78.24%. Further studies showed XZ34-1 could produce antifungal bioactive substances and had the potential of promoting plant growth. Lipopeptide genes detection with PCR indicated that strain XZ34-1 may produce lipopeptides. Furthermore, activities of defense-related enzymes were enhanced in wheat seedlings after inoculation with B.sorokiniana and treatment with CF, which showed induced resistance could be produced in wheat to resist pathogens. These results reveal that strain XZ34-1 is a promising candidate for application as a biological control agent against B.sorokiniana.

An integrated self-powered 3D printed sample concentrator for highly sensitive molecular detection of HIV in whole blood at the point of care.

Rapid and efficient biological sample preparation and pretreatment are crucial for highly sensitive, reliable and reproducible molecular detection of infectious diseases. Herein, we report a self-powered, integrated sample concentrator (SPISC) for rapid plasma separation, pathogen lysis, nucleic acid trapping and enrichment at the point of care. The proposed sample concentrator uses a combination of gravitational sedimentation of blood cells and capillary force for rapid, self-powered plasma separation. The pathogens (e.g., HIV virus) in separated plasma were directly lysed and pathogen nucleic acid was enriched by an integrated, flow-through FTA® membrane in the concentrator, enabling highly efficient nucleic acid preparation. The FTA® membrane of the SPISC is easy to store and transport at room temperature without need for uninterrupted cold chain, which is crucial for point of care sampling in resource-limited settings. The platform has been successfully applied to detect HIV virus in blood samples. Our experiments show that the sample concentrator can achieve a plasma separation efficiency as high as 95% and a detection sensitivity as low as 10 copies per 200 µL blood (∼100 copies per mL plasma) with variability less than 7%. The sample concentrator described is fully compatible with downstream nucleic acid detection and has great potential for early diagnostics, monitoring and management of infectious diseases at the point of care.

Effects of N-terminal modifications on the stability of antimicrobial peptide SAMP-A4 analogues against protease degradation.

Infections with multidrug-resistant (MDR) pathogens are increasingly concerning for public health. Synthesized antimicrobial peptide A4 (SAMP-A4), a peptide computationally designed by our research team, is a potential drug candidate. However, the antimicrobial peptide SAMP-A4 is easily degraded in serum. To obtain SAMP-A4 analogues with high biostability, chemical modifications at its N-terminus, including fatty acid conjugation, glycosylation and PEGylation, were carried out. The results showed that the introduction of hydrophobic fatty acids at the N-terminus of SAMP-A4 is better than hydrophilic glycosylation and PEGylation. With increasing fatty acid chain length, the stability of SAMP-A4 analogues in serum and trypsin solutions is significantly improved, and the activities against MDR bacteria and Candida are significantly enhanced. There is no obvious change in haemolysis even when hexanoic acid is coupled with SAMP-A4, so the resulting analogue SAMP-A4-C6, SAMP-A4 conjugated with hexanoic acid, is the most likely of the analogues to become a drug.

Primary research on sampling scheme for analyzing mycotoxin distribution in wheat and rice fields.

BACKGROUND: Mycotoxins are among the most severe food contaminants. Deoxynivalenol and aflatoxin contamination are predominant in wheat and rice, respectively. Nowadays, there are no standardized and approved grain-sampling schemes worldwide. This study aimed to develop a scientific grain-sampling scheme to investigate the regularity of mycotoxin distributed in wheat and rice fields. The data were analyzed with analysis of variance and cluster analysis to select a better sampling scheme. RESULTS: Considering the influences of the weather before harvest (temperature, humidity, wind direction, and other conditions), we sampled grains from different places in different farmlands and detected the mycotoxin content of the sampled grains. The mycotoxin content had extremely significant differences in the area of rice fields (P<0.01) and significant differences in the area of wheat fields (P<0.05). The filtering effect existed peripheral the field areas, especially peripheral the humid areas, where the fungi were filtered and the toxin were easily produced. Furthermore, the upwind direction peripheral the field areas cause more filterature effect than other wind direction. Although 97% of mycotoxins in wheat can be removed through the shelling process, the toxin content were not obviously affected by rice lodging in the field. According to the cluster analysis, the peripheral and middle areas were divided into the same group with higher mycotoxin content. CONCLUSION: This paper developed a sampling scheme to detect the mycotoxin content of wheat and rice in the field, considering the temperature and humidity of the weather, locations, and other grain contamination conditions before harvest. Meanwhile, the sampling rule of lodging and wind direction in the field was also assayed. © 2021 Society of Chemical Industry.

Antimicrobial peptide CGA-N12 decreases the Candida tropicalis mitochondrial membrane potential via mitochondrial permeability transition pore.

Amino acid sequence from 65th to 76th residue of the N-terminus of Chromogranin A (CGA-N12) is an antimicrobial peptide (AMP). Our previous studies showed that CGA-N12 reduces Candida tropicalis mitochondrial membrane potential. Here, we explored the mechanism that CGA-N12 collapsed the mitochondrial membrane potential by investigations of its action on the mitochondrial permeability transition pore (mPTP) complex of C. tropicalis. The results showed that CGA-N12 induced cytochrome c (Cyt c) leakage, mitochondria swelling and led to polyethylene glycol (PEG) of molecular weight 1000 Da penetrate mitochondria. mPTP opening inhibitors bongkrekic acid (BA) could contract the mitochondrial swelling induced by CGA-N12, but cyclosporin A (CsA) could not. Therefore, we speculated that CGA-N12 could induce C. tropicolis mPTP opening by preventing the matrix-facing (m) conformation of adenine nucleotide transporter (ANT), thereby increasing the permeability of the mitochondrial membrane and resulted in the mitochondrial potential dissipation.

RgC3H Involves in the Biosynthesis of Allelopathic Phenolic Acids and Alters Their Release Amount in Rehmannia glutinosa Roots.

Rehmannia glutinosa production is affected by replanting disease, in which autotoxic harm to plants is mediated by endogenous phenolic acids as allelopathic compounds found in root exudates. These phenolic acids are mostly phenylpropanoid products of plants' secondary metabolisms. The molecular mechanism of their biosynthesis and release has not been explored in R. glutinosa. P-coumarate-3-hydroxylase (C3H) is the second hydroxylase gene involved in the phenolic acid/phenylpropanoid biosynthesis pathways. C3Hs have been functionally characterized in several plants. However, limited information is available on the C3H gene in R. glutinosa. Here, we identified a putative RgC3H gene and predicted its potential function by in silico analysis and subcellular localization. Overexpression or repression of RgC3H in the transgenic R. glutinosa roots indicated that the gene was involved in allelopathic phenolic biosynthesis. Moreover, we found that these phenolic acid release amount of the transgenic R. glutinosa roots were altered, implying that RgC3H positively promotes their release via the molecular networks of the activated phenolic acid/phenylpropanoid pathways. This study revealed that RgC3H plays roles in the biosynthesis and release of allelopathic phenolic acids in R. glutinosa roots, laying a basis for further clarifying the molecular mechanism of the replanting disease development.

Effects of CGA-N12 on the membrane structure of Candida tropicalis cells.

The antimicrobial peptide CGA-N12 (NH2-ALQGAKERAHQQ-COOH) is an active peptide derived from chromogranin A (CGA) and consists of the 65th to 76th amino acids of the N-terminus. The results of our previous studies showed that CGA-N12 exerts anti-Candida activity by inducing apoptosis without destroying the integrity of cell membranes. In this study, the effect of CGA-N12 on the cell membrane structure of Candida tropicalis was investigated. CGA-N12 resulted in the dissipation of the membrane potential, the increase in membrane fluidity, and the outflow of potassium ions in C. tropicalis without significantly changing the ergosterol level. Fluorescence quenching was applied to evaluate the membrane channel characteristics induced by CGA-N12 through detection of the following: membrane permeability of hydrated Cl- (ϕ ≈ 0.66 nm) using the membrane-impermeable halogen anion-selective fluorescent dye lucigenin, passage of the membrane-impermeable dye carboxyfluorescein (CF) (ϕ ≈ 1 nm) through the membrane, and membrane permeation of H3O+ based on the membrane non-permeable pH-sensitive fluorescent dye 8-hydroxypyrene-1,3,6-trisulfonic acid, trisodium salt (HPTS). In conclusion, CGA-N12 can induce the formation of non-selective ion channels <1 nm in diameter in the membranes of C. tropicalis, resulting in the leakage of potassium ions, chloride ions, and protons, among others, leading to dissipation of the membrane potential. As a result, the fluidity of membranes is increased without destroying the synthesis of ergosterol is not affected.

Upregulation of miR-376c-3p alleviates oxygen-glucose deprivation-induced cell injury by targeting ING5.

BACKGROUND: The expression level of miR-376c-3p is significantly lower in infants with neonatal hypoxic-ischemic encephalopathy (HIE) than in healthy infants. However, the biological function of this microRNA remains largely elusive. METHODS: We used PC-12 and SH-SY5Y cells to establish an oxygen-glucose deprivation (OGD) cell injury model to mimic HIE in vitro. The miR-376c-3p expression levels were measured using quantitative reverse transcription PCR. The CCK-8 assay and flow cytometry were utilized to evaluate OGD-induced cell injury. The association between miR-376c-3p and inhibitor of growth 5 (ING5) was validated using the luciferase reporter assay. Western blotting was conducted to determine the protein expression of CDK4, cyclin D1, Bcl-2 and Bax. RESULTS: MiR-376c-3p was significantly downregulated in the OGD-induced cell injury model. Its overexpression elevated cell viability and impaired cell cycle G0/G1 phase arrest and apoptosis in PC-12 and SH-SY5Y cells after OGD. Downregulation of miR-376c-3p gave the opposite results. We further demonstrated that ING5 was a negatively regulated target gene of miR-376c-3p. Importantly, ING5 knockdown had a similar effect to miR-376c-3p-mediated protective effects against cell injury induced by OGD. Its overexpression abolished these protective effects. CONCLUSION: Our data suggest that miR-376c-3p downregulated ING5 to exert protective effects against OGD-induced cell injury in PC-12 and SH-SY5Y cells. This might represent a novel therapeutic approach for neonatal HIE treatment.

Loss of CXCR6 coreceptor usage characterizes pathogenic lentiviruses.

Pandemic HIV-1 originated from the cross-species transmission of SIVcpz, which infects chimpanzees, while SIVcpz itself emerged following the cross-species transmission and recombination of monkey SIVs, with env contributed by the SIVgsn/mus/mon lineage that infects greater spot-nosed, mustached and mona monkeys. SIVcpz and HIV-1 are pathogenic in their respective hosts, while the phenotype of their SIVgsn/mus/mon ancestors is unknown. However, two well-studied SIV infected natural hosts, sooty mangabeys (SMs) and African green monkeys (AGMs), typically remain healthy despite high viral loads; these species express low levels of the canonical coreceptor CCR5, and recent work shows that CXCR6 is a major coreceptor for SIV in these hosts. It is not known what coreceptors were used by the precursors of SIVcpz, whether coreceptor use changed during emergence of the SIVcpz/HIV-1 lineage, and what T cell subsets express CXCR6 in natural hosts. Using species-matched coreceptors and CD4, we show here that SIVcpz uses only CCR5 for entry and, like HIV-1, cannot use CXCR6. In contrast, SIVmus efficiently uses both CXCR6 and CCR5. Coreceptor selectivity was determined by Env, with CXCR6 use abrogated by Pro326 in the V3 crown, which is absent in monkey SIVs but highly conserved in SIVcpz/HIV-1. To characterize which cells express CXCR6, we generated a novel antibody that recognizes CXCR6 of multiple primate species. Testing lymphocytes from SM, the best-studied natural host, we found that CXCR6 is restricted to CD4+ effector memory cells, and is expressed by a sub-population distinct from those expressing CCR5. Thus, efficient CXCR6 use, previously identified in SM and AGM infection, also characterizes a member of the SIV lineage that gave rise to SIVcpz/HIV-1. Loss of CXCR6 usage by SIVcpz may have altered its cell tropism, shifting virus from CXCR6-expressing cells that may support replication without disrupting immune function or homeostasis, towards CCR5-expressing cells with pathogenic consequences.

CGA-N12, a peptide derived from chromogranin A, promotes apoptosis of Candida tropicalis by attenuating mitochondrial functions.

CGA-N12 (the amino acid sequence from the 65th to the 76th residue of the N-terminus of chromagranin A) is an antifungal peptide derived from human chromogranin A (CGA). In our previous investigation, CGA-N12 was found to have specific anti-candidal activity, though the mechanism of action remained unclear. Here, we investigated the effects of CGA-N12 on mitochondria. We found that CGA-N12 induced an over-generation of intracellular reactive oxygen species and dissipation in mitochondrial membrane potential, in which the former plays key roles in the initiation of apoptosis and the latter is a sign of the cell apoptosis. Accordingly, we assessed the apoptosis features of Candida tropicalis cells after treatment with CGA-N12 and found the following: leakage of cytochrome c and uptake of calcium ions into mitochondria and the cytosol; metacaspase activation; and apoptotic phenotypes, such as chromatin condensation and DNA degradation. In conclusion, CGA-N12 is capable of inducing apoptosis in C. tropicalis cells through mitochondrial dysfunction and metacaspase activation. Antifungal peptide CGA-N12 from human CGA exhibits a novel apoptotic mechanism as an antifungal agent.