Identification of phytochemicals in Qingfei Paidu decoction for the treatment of coronavirus disease 2019 by targeting the virus-host interactome.

Qingfei Paidu decoction (QFPDD) has been clinically proven to be effective in the treatment of coronavirus disease 2019 (COVID-19). However, the bioactive components and therapeutic mechanisms remain unclear. This study aimed to explore the effective components and underlying mechanisms of QFPDD in the treatment of COVID-19 by targeting the virus-host interactome and verifying the antiviral activities of its active components in vitro. Key active components and targets were identified by analysing the topological features of a compound-target-pathway-disease regulatory network of QFPDD for the treatment of COVID-19. The antiviral activity of the active components was determined by a live virus infection assay, and possible mechanisms were analysed by pseudotyped virus infection and molecular docking assays. The inhibitory effects of the components tested on the virus-induced release of IL-6, IL-1ß and CXCL-10 were detected by ELISA. Three components of QFPDD, oroxylin A, hesperetin and scutellarin, exhibited potent antiviral activities against live SARS-CoV-2 virus and HCoV-OC43 virus with IC50 values ranging from 18.68 to 63.27 µM. Oroxylin A inhibited the entry of SARS-CoV-2 pseudovirus into target cells and inhibited SARS-CoV-2 S protein-mediated cell-cell fusion by binding with the ACE2 receptor. The active components of QFPDD obviously inhibited the IL-6, IL-1ß and CXCL-10 release induced by the SARS-CoV-2 S protein. This study supports the clinical application of QFPDD and provides an effective analysis method for the in-depth study of the mechanisms of traditional Chinese medicine (TCM) in the prevention and treatment of COVID-19.

Molecular characterization and antiparasitic activity analysis of a novel piscidin 5-like type 4 from Larimichthys crocea.

Cryptocaryon irritans is an obligate parasitic ciliate protozoan that can infect various commercially important mariculture teleosts and cause high lethality and economic loss, especially Larimichthys crocea. Current methods of controlling or preventing this parasite with chemicals or antibiotics are widely considered to be environmentally harmful. The antiparasitic activity of some antimicrobial peptides (AMPs) attracted extensive attention of scholars. In the study, a novel piscidin 5-like type 4 (termed Lc-P5L4) excavated from comparative transcriptome of C. irritans - immuned L. crocea was identified and characterized. Sequence analysis shows the full-length cDNA of Lc-P5L4 is 539 bp containing an open reading frame (ORF) of 198 bp which encodes a peptide of 65 amino acid residues. The genome consists of three exons and two introns which exist in its ORF, and all the exon-intron boundaries are in accordance with classical GT-AG rule (GT/intron/AG). Multiple alignments indicate the signal peptides share highly conserved identity, while mature peptides are more diverse. Phylogenetic analysis displays Lc-P5L4 clusters together with other members of piscidin 5-like family. Next, quantitative Real-time PCR (qRT-PCR) detection found C. irritans infection could upregulate Lc-P5L4 expression level in all tested tissues significantly, it appeared earliest upregulation in the theronts infection stage in the head kidney; the expression contents reached to maximum level in the intestine, gill and muscle during trophonts falling off stage; while it was just upregulated during secondary bacterial infection stage in the liver and spleen. The data showed Lc-P5L4 upregulation time points were in accordance with different infection stages. With recombinant Lc-P5L4 (rLc-P5L4) obtained through Escherichia coli system, in vitro assay showed rLc-P5L4 could cause cilia deactivation, cell bodiesclumping and sticking to each other, then cell membrane rupture and contents leakage. The data illustrated Lc-P5L4 played critical roles in the immune defense against C. irritans infection, and provided another proof that piscidins exhibit multiple anti- C. irritans features.

Trilobatin as an HIV-1 entry inhibitor targeting the HIV-1 Gp41 envelope.

HIV-1 transmembrane protein gp41 plays a crucial role by forming a stable six-helix bundle during HIV entry. Due to highly conserved sequence of gp41, the development of an effective and safe small-molecule compound targeting gp41 is a good choice. Currently, natural polyanionic ingredients with anti-HIV activities have aroused concern. Here, we first discovered that a glycosylated dihydrochalcone, trilobatin, exhibited broad anti-HIV-1 activity and low cytotoxicity in vitro. Site-directed mutagenesis analysis suggested that the hydrophobic residue (I564) located in gp41 pocket-forming site is pivotal for anti-HIV activity of trilobatin. Furthermore, trilobatin displayed synergistic anti-HIV activities combined with other antiretroviral agents. Trilobatin has a good potential to be developed as a small-molecule HIV-1 entry inhibitor for clinical combination therapy.

The effect of chromium picolinate supplementation on the pancreas and macroangiopathy in type II diabetes mellitus rats.

PURPOSE: The aim was to explore the effect of the chromium picolinate (CrPic) administration on the pancreas and macroangiopathy of type II diabetes mellitus rats. METHODS: The type II diabetes mellitus (T2DM) rat model was induced by low-dose streptozotocin (STZ). The rats were randomly divided into 5 groups (ten rats in each group). After supplementing CrPic for 15 weeks, the histopathological examination was performed by hematoxylin-eosin (HE) staining. Serum insulin and NO level were determined by radioimmunoassay and colorimetry, respectively. Serum glycosylated hemoglobin (HbA1C), adiponectin (APN), advanced glycation end products (AGES), and apelin were measured by ELISA. Real-time reverse transcription polymerase chain reaction (RT-PCR) was applied for detecting the mRNA expression of APN and apelin. RESULTS: After CrPic treatment, compared with the T2DM control group (group 2), pancreas sections stained with HE showed the completed pancreatic cells structure and no inflammatory infiltration in groups 4 and 5. In addition, the levels of serum NO and insulin were significantly increased and the serum levels of HbA1C, AGES, APN, and apelin were significantly decreased in groups 4 and 5 compared with group 2. The mRNA expression of APN and apelin in groups 4 and 5 was also recovered to the normal level. CONCLUSION: CrPic can recover the function of Β-cells and alleviate macroangiopathy in STZ-induced T2DM rats.

[Virtual screening of small molecular HIV-1 entry inhibitor NC-2 targeting gp120 and its action mechanism].

OBJECTIVE: To screen the HIV-1 entry inhibitors targeting HIV-1 gp120 from the IBS natural product database by virtual screening based on the binding mode of the neutralizing antibody VRC01 with HIV-1 gp120 and investigate the anti-viral activities of the inhibitors and their action mechanisms. METHODS: The binding interaction of the candidate molecules binding gp120 and changes of the binding free energy were analyzed by MM-PBSA calculation. The anti-HIV-1 activities of the tested compounds were detected by HIV-1 pseudotyped virus, laboratory-adapted HIV-1 and a cell-cell fusion assay. The cytotoxicity of the studied molecules was examined by XTT colorimetric assay. The mechanisms of the anti-viral activities of the candidate molecules were analyzed using enzyme-linked immunosorbent assay. RESULTS: A total of 19 molecules with distinct reduction of the binding free energy after binding with gp120 were screened from 40000 molecules. Among them, NC-2 showed anti-HIV-1 activities against HIV-1 pseudotyped virus and laboratory-adapted HIV-1, and was capable of blocking HIV-1 envelope-mediated cell-cell fusion. The IC50 of NC-2 for inhibiting HIV-1IIIB and pseudotyped HIV-1JRFL infection were 1.95∓0.44 µmol/L and 10.58∓0.13 µmol/L, respectively. The results of ELISA suggested that NC-2 could inhibit the binding of HIV-1 gp120 to CD4 without blocking the formation of gp41 six-helix bundle in vitro. CONCLUSION: This computer-based virtual screening method can be used to screen HIV-1 entry inhibitors targeting gp120. Using this virtual screening approach combined with anti-viral activity screening, we obtained a potent HIV-1 entry inhibitor NC-2 with novel structure.

A natural theaflavins preparation inhibits HIV-1 infection by targeting the entry step: potential applications for preventing HIV-1 infection.

Theaflavins are the major components of tea polyphenols in brewed black tea. We previously reported that theaflavin derivatives, such as TF3, inhibited HIV-1 entry by targeting gp41. However, it is difficult to purify the individual theaflavins and the purified compounds are highly unstable. To develop theaflavins as affordable anti-HIV-1 microbide for preventing HIV sexual transmission, we intended to use an economic natural preparation containing 90% of theaflavins (TFmix). Its antiviral activity against HIV-1 strains was evaluated in vitro using p24 production and luciferase assays. The mechanism by which TFmix inhibits HIV-1 infection was investigated using time-of-addition, cell-cell fusion and biophysical assays. The data suggested TFmix exhibited potent anti-HIV-1 activity on lab-adapted and primary HIV-1 strains with IC(50) less than 1.20 µM. It also effectively inhibited infection by T-20 resistant HIV-1 strains. The mechanism studies suggest that TFmix mainly inhibit the HIV-1 entry by targeting gp41 since it is effective in inhibiting gp41 six-helix bundle (6-HB) formation and HIV-1 envelope protein-mediated cell-cell fusion. TFmix could also inhibit HIV-1 reverse transcriptase (RT) activity, but the IC(50) is about 8-fold higher than that for inhibiting gp41 6-HB formation, suggesting RT is not a major target for TFmix. In conclusion, TFmix is an economic natural product preparation containing high content of theaflavins with potent anti-HIV-1 activity by targeting the viral entry step through the disruption of gp41 6-HB core structure. It has a potential to be developed as a safe and affordable topical microbicide for preventing sexual transmission of HIV.