Disrupted establishment of anaerobe and facultative anaerobe balance in preterm infants with extrauterine growth restriction.

Background: Extrauterine growth restriction (EUGR) in preterm birth infants could have long-term adverse impacts on health. Less is known about the gut microbiota regarding its establishment in early life and its role in long-term growth in preterm birth infants. Methods: A prospective, longitudinal observational study was conducted with 67 preterm infants in a level III neonatal intensive care unit. Clinical information was obtained from medical records, and fecal samples were collected weekly during hospitalization and processed for 16S rRNA gene sequencing. Results: The bacterial profiles from the weekly sampling of preterm infants demonstrated that the early-life gut microbiota was clustered into the following four stages in chronological order: stage 1: 0-4 days, stage 2: 1-2 weeks, stage 3: 3-7 weeks, and stage 4: 8-10 weeks. The development of gut microbiota showed latency at stage 4 in EUGR infants compared with that in non-EUGR infants, which resulted from their consistently high level of facultative anaerobes, including Enterobacteriaceae and Staphylococcus, and lack of obligate anaerobes, including Clostridium and Veillonella. In the 2-year follow-up, infants with a high level of obligate anaerobes-to-facultative anaerobes ratio at stage 4 had a lower risk of long-term growth restriction at the margin of statistical significance. Conclusion: The results of this study indicate that the development of gut microbiota in the early life of EUGR infants is delayed compared with that of non-EUGR infants. The obligate-to-facultative anaerobes ratio could be an indicator of the maturity of gut microbiota development and associated with the risk of long-term growth restriction in preterm infants.

Clinical features and recurrence of Corynebacterium kroppenstedtii infection in patients with mastitis.

BACKGROUND: Few studies have investigated the differences in clinical features of patients with mastitis following Corynebacterium kroppenstedtii infection, and most focused on the bacterial antimicrobial susceptibility, detection methods and therapy. METHODOLOGY: There were 133 patients with mastitis infected by C. kroppenstedtii between August 2016 and September 2019. C. kroppenstedtii was identified using mass spectrometry. The demographics, clinical diagnosis, laboratory test results of different types of mastitis combined with bacillus infection, and the effects of different treatments in reducing recurrence were compared. RESULTS: The incidence of pus following C. kroppenstedtii infection was higher in patients with non-granulomatous lobular mastitis (NGLM; 56.6%) than in those with granulomatous lobular mastitis (GLM; 33.3%; χ2 = 7.072, p = 0.008). While C-reactive protein (CRP) was higher in the GLM group (12.50 mg/L) than in the NGLM group (6.05 mg/L; Z = - 2.187, p = 0.029). Treatment with local lavage (triamcinolone acetonide) and antibiotics (cefuroxime) showed a recurrent rate of 25.9% in C. kroppenstedtii infection. CONCLUSION: Increased pus, large masses, and an elevated CRP level may occur in patients with mastitis infected by C.kroppenstedtii. These clinical features may guide the determination of the bacterial infection in patients with mastitis. Combining an antibiotic with a triamcinolone acetonide lavage, preferably cefuroxime, may reduce the recurrence.

Elimination of Grain Boundary Defects in Zeolitic Imidazolate Framework ZIF-95 Membrane via Solvent-Free Secondary Growth.

Metal-organic framework membranes are usually prepared by in situ or secondary growth in a solution/hydrogel. The use of organic solvents may cause safety and environmental problems and produce solvent-induced defects. Here, highly oriented and permselective ZIF-95 membranes are prepared for the first time via a solvent-free secondary growth method. The solvent-free growth is not only helpful to control the membrane microstructure and thickness, but also to reduce the intercrystalline defects. In case of solvent-free growth, a perfectly oriented structure leads to an outstanding reduction of intercrystalline defects and transport resistances. For the separation of equimolar binary gas mixtures by using the highly oriented ZIF-95 membrane at 25 °C and 1 bar, the mixture separation factors of H2 /CO2 and H2 /CH4 are 184 and 140, respectively, with H2 permeance of over 1.9×10-7  mol m-2 s-1 Pa-1 which are much higher than those of the randomly oriented ZIF-95 membrane.

Oral ibuprofen promoted cholestatic liver disease in very low birth weight infants with patent ductus arteriosus.

OBJECTIVES: Hemodynamically significant patent ductus arteriosus (hsPDA) in very low birth weight (VLBW) infants is routinely treated in many countries with oral ibuprofen. This study retrospectively assessed whether the risk of cholestatic liver disease (CLD) increased due to oral ibuprofen administration in VLBW infants. METHODS: A total of 122 VLBW preterm infants (26∼32 weeks, birth weight<1500g) diagnosed with patent ductus arteriosus (PDA) admitted to our neonatal intensive care unit (NICU) between September 2016 to August 2018 were included. Sixty-four infants were diagnosed with hs-PDA and received ibuprofen treatment. VLBW infants with PDA untreated with ibuprofen served as controls. Soybean oil and fat emulsions were routinely added to parenteral nutrition (PN). Once CLD was diagnosed, the fat emulsions were immediately replaced with multi-oil fat emulsion injections. To assess the independent association of treatment and duration of ibuprofen with CLD and duration of fasting and PN, binary logistic regression or multivariate linear regression analyses were conducted, adjusting for major confounders (birth weight, gestational age, Clinical Risk Index for Babies, and cholestasis-associated risk factors). RESULTS: The duration of PN increased due to ibuprofen treatment for 6.559 days (95% CI: 1.769, 11.349; P=0.008), and the risk of prolonged fasting (cutoff>5 days) might have increased due to ibuprofen treatment (OR: 3.043, 95% CI: 0.965, 9.594; P=0.057). Furthermore, CLD was influenced by ibuprofen treatment (OR: 6.730; 95% CI: 1.279, 35.41; P=0.024), early thrombocytopenia 7 days postnatal (OR: 6.996; 95% CI: 1.769, 27.658; P=0.004), and late onset sepsis (OR: 6.976; 95% CI: 1.561, 31.169; P=0.011). Further analysis adjusting for cholestasis-associated risk factors revealed that CLD was influenced by the duration of ibuprofen treatment (OR: 2.864; 95% CI: 1.104, 7.422; P=0.030), Platlets counts 7 days postnatal (OR: 0.971; 95% CI: 0.950, 0.994; P=0.013), and duration of antibiotics (OR: 1.134; 95% CI: 1.002, 1.282; P=0.046). CONCLUSIONS: This retrospective study indicated oral ibuprofen duration-dependently increased the risk of CLD in VLBW infants with PDA, and early thrombocytopenia served as the critical risk factor.

An oligothiophene compound neutralized influenza A viruses by interfering with hemagglutinin.

BACKGROUND AND OBJECTIVES: Recently influenza pandemic outbreaks were caused by emerging H5N1, H7N9 and H1N1 viruses. However, virucidal disinfectants are mainly unspecific and toxic. It is tactical to discover specific virucidal compounds. METHODS: The inhibitory potency was determined in H5N1 pseudovirus system; Interactions of compounds with hemagglutinin (HA) were detected with surface plasmon resonance (SPR) and further calculated with molecular docking. Virucidal effect was also estimated in influenza virus A/Puerto Rico/8/34(H1N1). Prevention efficacy was further estimated in mice model. RESULTS: Oligothiophene compound 4sc was potently virucidal against H5N1 pseudovirus with selective index>1169 (IC50=0.17±0.01µM). Pseudovirus assay revealed 4sc may interact with HA. However, HA inhibition test indicated 4sc did not interact with receptor pocket in HA. SPR detection revealed 4sc interacted directly with HA and its HA2 subunits. Molecular docking analysis revealed that 4sc interacted with the cavity of HA2 stem region and HA1-HA2 interface which consist of 7 residues: L22, K262, G472 and F1102 in HA2; M241, E251 and N271 in HA1. 4sc also potently and irreversibly neutralized PR8 (H1N1) virus, causing 105.06±0.26 fold decrease of virus titer after exposure for 10min. 4sc blocked PR8 transmission to MDCK cells. Amazingly, virucidal effect of 4sc was not significantly reduced even at 4°C. Furthermore, 4sc blocked viral transmission to mice. CONCLUSION: Oligothiophene compound 4sc is a novel selective virucide of influenza virus, which blocks entry by interfering viral hemagglutinin. Due to promising safety profile and stable virucidal effect at 4°C, 4sc may be useful in disinfecting H5N1 and H1N1 influenza virus.

Structure-activity relationships of 3-O-ß-chacotriosyl oleanic acid derivatives as entry inhibitors for highly pathogenic H5N1 influenza virus.

Highly pathogenic H5N1 virus (H5N1) entry is a key target for the development of novel anti-influenza agents with new mechanisms of action. In our continuing efforts to identify novel potential anti-H5N1 entry inhibitors, a series of 3-O-ß-chacotriosyl oleanolic acid analogs have been designed, synthesized and evaluated as H5N1 entry inhibitors based on two small molecule inhibitors 1 and 2 previously discovered by us. The anti-H5N1 entry activities were determined based on HA/HIV and VSVG/HIV entry assays. Compound 15 displayed the most promising anti-H5N1 entry activities with average IC50 values of 4.05µM and good selective index (22.9). Detailed structure-activity relationships (SARs) studies suggested that either the introduction of an additional oxo group to position 11 at OA or alteration of the C-3 configuration of OA from 3ß- to 3α-forms can significantly enhance the selective index while maintaining their antiviral activities in vitro. Molecular simulation analysis confirmed that the compounds exert their inhibitory activity through binding tightly to hemagglutinin (HA2) protein near the fusion peptide and prevent virus entry.

Heat Shock Factor 1 Mediates Latent HIV Reactivation.

HSF1, a conserved heat shock factor, has emerged as a key regulator of mammalian transcription in response to cellular metabolic status and stress. To our knowledge, it is not known whether HSF1 regulates viral transcription, particularly HIV-1 and its latent form. Here we reveal that HSF1 extensively participates in HIV transcription and is critical for HIV latent reactivation. Mode of action studies demonstrated that HSF1 binds to the HIV 5'-LTR to reactivate viral transcription and recruits a family of closely related multi-subunit complexes, including p300 and p-TEFb. And HSF1 recruits p300 for self-acetylation is also a committed step. The knockout of HSF1 impaired HIV transcription, whereas the conditional over-expression of HSF1 improved that. These findings demonstrate that HSF1 positively regulates the transcription of latent HIV, suggesting that it might be an important target for different therapeutic strategies aimed at a cure for HIV/AIDS.

Structure-activity relationships of 3-O-ß-chacotriosyl oleanane-type triterpenoids as potential H5N1 entry inhibitors.

A series of 3-O-ß-chacotriosyl oleanolic acid analogs have been designed, synthesized and evaluated as H5N1 entry inhibitors based on a small molecule inhibitor saponin 1 previously discovered by us. Detailed structure-activity relationships (SARs) studies on the aglycone of compound 1 indicated that the subtle modification of oleanolic acid as an aglycon has key influences on the antiviral activity. These results suggested that either the introduction of a disubstituted amide structure at the 17-COOH of OA or alteration of the C-3 configuration of OA from 3ß-to 3α-forms can significantly improve the selective index while maintaining their antiviral activities in vitro. Compound 8 was selected for further mechanistic study because of its distinguished inhibition activity and good selective index. Molecular simulation study and surface plasmon resonance analysis confirmed that compound 8 stabilized HA2 subunit of hemagglutinin (HA) by binding with amino acid residues LYS-26, ASN-53, ASN-27 and ASN-50, therefore may prevent HA from conformational rearranging, which is a critical step for viral entry.

New influenza A Virus Entry Inhibitors Derived from the Viral Fusion Peptides.

Influenza A viral (IAV) fusion peptides are known for their important role in viral-cell fusion process and membrane destabilization potential which are compatible with those of antimicrobial peptides. Thus, by replacing the negatively or neutrally charged residues of FPs with positively charged lysines, we synthesized several potent antimicrobial peptides derived from the fusogenic peptides (FPs) of hemagglutinin glycoproteins (HAs) of IAV. The biological screening identified that in addition to the potent antibacterial activities, these positively charged fusion peptides (pFPs) effectively inhibited the replication of influenza A viruses including oseltamivir-resistant strain. By employing pseudovirus-based entry inhibition assays including H5N1 influenza A virus (IAV), and VSV-G, the mechanism study indicated that the antiviral activity may be associated with the interactions between the HA2 subunit and pFP, of which, the nascent pFP exerted a strong effect to interrupt the conformational changes of HA2, thereby blocking the entry of viruses into host cells. In addition to providing new peptide "entry blockers", these data also demonstrate a useful strategy in designing potent antibacterial agents, as well as effective viral entry inhibitors. It would be meaningful in treatment of bacterial co-infection during influenza pandemic periods, as well as in our current war against those emerging pathogenic microorganisms such as IAV and HIV.

Super short membrane-active lipopeptides inhibiting the entry of influenza A virus.

Influenza A viruses (IAV) are significant pathogens that result in millions of human infections and impose a substantial health and economic burdens worldwide. Due to the limited anti-influenza A therapeutics available and the emergence of drug resistant viral strains, it is imperative to develop potent anti-IAV agents with different mode of action. In this study, by applying a pseudovirus based screening approach, two super short membrane-active lipopeptides of C12-KKWK and C12-OOWO were identified as effective anti-IAV agents with IC50 value of 7.30±1.57 and 8.48±0.74 mg/L against A/Puerto Rico/8/34 strain, and 6.14±1.45 and 7.22±0.67 mg/L against A/Aichi/2/68 strain, respectively. The mechanism study indicated that the anti-IAV activity of these peptides would result from the inhibition of virus entry by interacting with HA2 subunit of hemagglutinin (HA). Thus, these peptides may have potentials as lead peptides for the development of new anti-IAV therapeutics to block the entry of virus into host cells.

[3-O-ß-chacotriosyl benzyl ursolate inhibits entry of H5N1 influenza virus into target cells].

OBJECTIVE: To study the inhibitory activities of 3-O-ß-chacotriosyl benzyl ursolate and its derivatives as potential new anti-influenza virus agents against the entry of H5N1 influenza viruses into the target cells. METHODS: Four target compounds were designed and synthesized, which were structurally related to the lead compound 3-O-ß-chacotriosyl methyl ursolate (1). The inhibitory activities of these compounds were tested at a cellular level psuedovirus system targeting H5N1 influenza viruse entry. RESULTS AND CONCLUSION: The compounds 1b, 1c and 1d showed potent inhibitory activities against the entry of A/Thailand/Kan353/2004 pseudovirus into the target cells, and among them compound 1d showed the strongest inhibitory activity with an IC50 value of 0.96 ± 0.10 µmol/L. The structure-activity relationship analysis of these compounds indicated that when 17-COOH of ursolic acid was esterified, introduction of Me groups rather than aryl groups more strongly enhanced the inhibitory activity. Changing 17-COOH of ursolic acid into amide could increase the antiviral activity and decrease the cytotoxicity of the compounds in MDCK cells.

Structure-activity relationships of 3-O-ß-chacotriosyl ursolic acid derivatives as novel H5N1 entry inhibitors.

A series of methyl ursolate 3-O-ß-chacotrioside analogs have been designed, synthesized and evaluated as H5N1 entry inhibitors based on a small molecule inhibitor saponin 3 previously discovered by us. Detailed structure-activity relationships (SARs) studies on the aglycone of compound 3 indicated that both the type of pentacyclic triterpene and the subtle modification of ursolic acid as an aglycon had key influences on the antiviral activity. These results suggested that either the introduction of a disubstituted amide structure at the 17-COOH of ursolic acid or alteration of the C-3 configuration of ursolic acid from 3ß-to 3α-forms was helpful to significantly improve the selective index while keeping their antiviral activities.

Bioactive metabolites produced by the endophytic fungus Phomopsis sp. YM355364.

A new compound, 16-acetoxycytosporone B (1), along with four known ones, dankasterone A (2), dankasterone B (3), 3beta,5alpha,9alpha-trihydroxy-(22E,24R)-ergosta-7,22-dien-6-one (4), and cyclonerodiol oxide (5), were isolated from Phomopsis sp. YM355364, an endophytic fungus of Aconitum carmichaeli. Their structures were characterized by spectral analysis. Compound 2 exhibited significant inhibitory activity against influenza A/Thailand/Kan353/2004(H5N1) pseudovirus with all IC50 value of 3.56 microM. Compounds 1, 2, and 4 showed either moderate or weak antifungal activities against four pathogenic fungi.

Novel hemagglutinin-based influenza virus inhibitors.

Influenza virus has caused seasonal epidemics and worldwide pandemics, which caused tremendous loss of human lives and socioeconomics. Nowadays, only two classes of anti-influenza drugs, M2 ion channel inhibitors and neuraminidase inhibitors respectively, are used for prophylaxis and treatment of influenza virus infection. Unfortunately, influenza virus strains resistant to one or all of those drugs emerge frequently. Hemagglutinin (HA), the glycoprotein in influenza virus envelope, plays a critical role in viral binding, fusion and entry processes. Therefore, HA is a promising target for developing anti-influenza drugs, which block the initial entry step of viral life cycle. Here we reviewed recent understanding of conformational changes of HA in protein folding and fusion processes, and the discovery of HA-based influenza entry inhibitors, which may provide more choices for preventing and controlling potential pandemics caused by multi-resistant influenza viruses.

Influenza A virus entry inhibitors targeting the hemagglutinin.

Influenza A virus (IAV) has caused seasonal influenza epidemics and influenza pandemics, which resulted in serious threat to public health and socioeconomic impacts. Until now, only 5 drugs belong to two categories are used for prophylaxis and treatment of IAV infection. Hemagglutinin (HA), the envelope glycoprotein of IAV, plays a critical role in viral binding, fusion and entry. Therefore, HA is an attractive target for developing anti­IAV drugs to block the entry step of IAV infection. Here we reviewed the recent progress in the study of conformational changes of HA during viral fusion process and the development of HA-based IAV entry inhibitors, which may provide a new choice for controlling future influenza pandemics.

Endogenous hydrogen sulfide mediates the cardioprotection induced by ischemic postconditioning in the early reperfusion phase.

Hydrogen sulfide (H(2)S), produced by cystanthionine-γ-lysase (CSE) in the cardiovascular system, has been suggested to be the third gasotransmitter in addition to nitric oxide (NO) and carbon monoxide (CO). The present study aimed to investigate the role of H(2)S in ischemic postconditioning (IPO) during the early period of reperfusion. IPO with 6 episodes of 10 sec reperfusion followed by 6 episodes of 10 sec ischemia (IPO 2') was administered when reperfusion was initiated. Cardiodynamics and the concentration of H(2)S were measured at 1, 2, 3, 4, 5, 10, 20, 30, 60, 90 and 120 min of reperfusion. Lactate dehydrogenase (LDH) levels and infarct size were determined at the end of the reperfusion. The concentration of H(2)S was stable during the whole experiment in the control group, whereas it reached a peak at the first minute of reperfusion in the ischemia-reperfusion (IR) group. The concentration of H(2)S at the first minute of reperfusion in the IPO 2' group was higher compared to that of the IR group, which correlated with cardioprotection including improved heart contractile function and reduced infarct size and LDH levels. However, the above effects of IPO 2' were attenuated by pre-treatment with blockade of endogenous H(2)S production with DL-propargylglycine for 20 min prior to global ischemia. Furthermore, we found that other forms of IPO, IPO commencing at 1 min after reperfusion (delayed IPO) or lasting only for 1 min (IPO 1'), failed to increase the concentration of H(2)S and protect the myocardium. We conclude that the peak of endogenous H(2)S in the early reperfusion phase is the key to cardioprotection induced by IPO.

Inhibition of hydrogen sulfide generation contributes to 1-methy-4-phenylpyridinium ion-induced neurotoxicity.

Reactive oxygen species (ROS) overproduction contributes to the neurotoxicity of 1-methy-4-phenylpyridinium ion (MPP(+)). Increasing studies have shown that hydrogen sulfide (H(2)S) is an endogenous antioxidant gas. We have hypothesized that MPP(+)-caused neurotoxicity may involve the imbalance of proportion to this endogenous protective antioxidant gas. The aim of this study is to evaluate whether MPP(+) disturbs H(2)S synthesis in PC12 cells, a clonal rat pheochromocytoma cell line, and whether disturbance of H(2)S generation induced by MPP(+) is an underlying mechanism of MPP(+)-induced neurotoxicity. We show that exposure of PC12 cells to MPP(+) causes a significant decrease in H(2)S generation and results in remarkable cell damage. We find that cystathionine-ß-synthetase (CBS) is catalyzed in PC12 cells to generate H(2)S, and that both expression and activity of CBS are inhibited by MPP(+) treatment. Exposure of sodium hydrosulfide (NaHS), a donor of H(2)S, extenuates MPP(+)-induced cytotoxicity and ROS accumulation in PC12 cells, while inhibition of CBS by amino-oxyacetate (AOAA) exacerbates the effects of MPP(+). These results indicate that MPP(+) neurotoxicity involves reduction of H(2)S production, which is caused by inhibition of CBS. This study provides novel insights into cell death observed in neurodegenerative disease such as Parkinson's disease.

Inhibition of endogenous hydrogen sulfide generation is associated with homocysteine-induced neurotoxicity: role of ERK1/2 activation.

Both elevated homocysteine and decreased hydrogen sulfide (H(2)S) are observed in the brains of Alzheimer's disease (AD) patients. Reactive oxygen species (ROS) overproduction contributes to the neurotoxicity of homocysteine; however, H(2)S is an endogenous antioxidant gas. Therefore, the aim of this study was to investigate whether the imbalance of proportion to this endogenous protective antioxidant gas is involved in homocysteine-caused neurotoxicity. We show that homocysteine inhibits the generation of endogenous H(2)S and the expression and activity of cystathionine-ß-synthetase (CBS), the main enzyme responsible for the generation of H(2)S in PC12 cells. S-Adenosylmethionine, an activator of CBS, not only prevents homocysteine-induced inhibition of endogenous H(2)S production but also attenuates homocysteine-triggered cytotoxicity and accumulation of ROS. We find that activation of ERK1/2 occurs in homocysteine-treated PC12 cells and blockade of ERK1/2 with U0126 abolished the homocysteine-induced cytotoxicity and inhibitory effect on endogenous H(2)S generation. These results indicate that homocysteine neurotoxicity involves reduction of H(2)S production, which is caused by inhibition of CBS and mediated by activation of ERK1/2. Our study suggests a promising future of H(2)S-based therapies for neurodegenerative diseases such as AD.

Hydrogen sulfide antagonizes homocysteine-induced neurotoxicity in PC12 cells.

Hydrogen sulfide (H2S) has been shown to protect neurons against oxidative stress. Lower levels of H(2)S as well as accumulation of homocysteine (Hcy), a strong risk of Alzheimer's disease (AD), are reported in the brains of AD patients. The aim of present study is to explore the protection of H2S against Hcy-induced cytotoxicity and apoptosis and the molecular mechanisms underlying in PC12 cells. We show that sodium hydrosulfide (NaHS), a H2S donor, protects PC12 cells against Hcy-mediated cytotoxicity and apoptosis by preventing both the loss of mitochondrial membrane potential (MMP) and the increase in intracellular reactive oxygen species (ROS) induced by Hcy. NaHS not only promotes the expression of bcl-2, but also blocks the down-regulation of bcl-2 by Hcy. These results indicate that H2S protects neuronal cells against neurotoxicity of Hcy by preserving MMP and attenuating ROS accumulation through up-regulation of bcl-2 level. Our study suggests a promising future of H2S-based therapies for neurodegenerative diseases such as AD.