Ent-labdane-type diterpene glycosides obtained from Rubus chingii Hu and their inhibitory effects on PDE5A activity.

In this study, 16 new ent-labdane-type diterpene glycosides, designated as goshonosides J1-J16 (1-16), along with nine previously known diterpene glycosides (17-25) were extracted from the fruits of Rubus chingii Hu. The structures of goshonosides J1-J16 were elucidated using various analytical techniques, such as nuclear magnetic resonance, electron capture detector ECD, high-resolution electrospray ionization mass spectrometry HREIMS, single-crystal X-ray diffraction, and hydrolysis. Furthermore, the isolates' efficacy in inhibiting the activity of phosphodiesterase type 5 A was evaluated. Goshonosides J1, J2, and G effectively inhibited the activity of the aforementioned enzyme (IC50 values: 6.15 ± 1.76, 3.27 ± 0.65, and 9.61 ± 2.36 µM, respectively). Our findings highlight the remarkable structural diversity of bioactive compounds in R. chingii Hu and offer insights into the use of this shrub.

Bioactive compounds from Huashi Baidu decoction possess both antiviral and anti-inflammatory effects against COVID-19.

The coronavirus disease 2019 (COVID-19) pandemic is an ongoing global health concern, and effective antiviral reagents are urgently needed. Traditional Chinese medicine theory-driven natural drug research and development (TCMT-NDRD) is a feasible method to address this issue as the traditional Chinese medicine formulae have been shown effective in the treatment of COVID-19. Huashi Baidu decoction (Q-14) is a clinically approved formula for COVID-19 therapy with antiviral and anti-inflammatory effects. Here, an integrative pharmacological strategy was applied to identify the antiviral and anti-inflammatory bioactive compounds from Q-14. Overall, a total of 343 chemical compounds were initially characterized, and 60 prototype compounds in Q-14 were subsequently traced in plasma using ultrahigh-performance liquid chromatography with quadrupole time-of-flight mass spectrometry. Among the 60 compounds, six compounds (magnolol, glycyrrhisoflavone, licoisoflavone A, emodin, echinatin, and quercetin) were identified showing a dose-dependent inhibition effect on the SARS-CoV-2 infection, including two inhibitors (echinatin and quercetin) of the main protease (Mpro), as well as two inhibitors (glycyrrhisoflavone and licoisoflavone A) of the RNA-dependent RNA polymerase (RdRp). Meanwhile, three anti-inflammatory components, including licochalcone B, echinatin, and glycyrrhisoflavone, were identified in a SARS-CoV-2-infected inflammatory cell model. In addition, glycyrrhisoflavone and licoisoflavone A also displayed strong inhibitory activities against cAMP-specific 3',5'-cyclic phosphodiesterase 4 (PDE4). Crystal structures of PDE4 in complex with glycyrrhisoflavone or licoisoflavone A were determined at resolutions of 1.54 Å and 1.65 Å, respectively, and both compounds bind in the active site of PDE4 with similar interactions. These findings will greatly stimulate the study of TCMT-NDRD against COVID-19.

[Identification of Bacterial Flora and Metabolic Function of Sediments in Different Channels of Duliujian River Basin, Tianjin].

Bacterial communities are highly sensitive to environmental changes, but their metabolic functions may be convergent under similar ecological conditions. In order to test this environmental attribute of the bacterial community and verify the feasibility in using the bacterial metabolic data to divide functional units at the river basin scale, the surface sediments in three continuous spatial units of the main stream (MS), left tributary (LT), and right tributary (RT) of the Duliujian River basin were selected as the research objects. Therefore, 16S rDNA high-throughput sequencing and the PICRUSt platform were used to analyze the bacterial diversity and metabolic function of sediments in different units. The results showed that there were no significant differences for the Shannon and Simpson indices between the different river channels (P>0.05); however, the bacterial diversity indices of Chao1, ACE, Observed_species, and PD_whole_tree of LT and RT were significantly higher than those of the MS (P<0.05). Moreover, the dominant bacterial phyla were not significantly different among the different river channels. In the present study, a total of 41 metabolic pathways were predicted based on the KEGG database, among which 34 metabolic pathways such as membrane transport, amino acid metabolism, and carbohydrate metabolism were significantly different. The functional metabolic abundance was higher in MS than that in RT and LT (P<0.05); however, there was no significant difference between those of RT and LT (P>0.05). Except for exchangeable phosphorus and nitrate nitrogen, there were no significant differences in other physicochemical properties among the different river channels (P>0.05). Furthermore, pH and iron/aluminum-phosphorus were found to significantly affect the bacterial structure, and SOM and TN were found to significantly affect the bacterial metabolic function. Overall, the sediments of MS and tributaries (LT and RT) of the river were clearly divided into level-1 and level-2 groups, indicating that the metabolic abundance of the bacterial community can be used as the key indicator in the division of functional units at the river basin scale. This study provided a theoretical basis and technical support for ecological functional division and management in coastal river basins.

Allometric releases of nitrogen and phosphorus from sediments mediated by bacteria determines water eutrophication in coastal river basins of Bohai Bay.

Although the Chinese government has conducted much work in recent years to reduce land-based pollutant discharge, eutrophication continues to occur frequently in many rivers, estuaries, and coastal waters. This may indicate that sediment is a major source rather than a sink for nitrogen (N) and phosphorus (P). To clarify the endogenous mechanisms of eutrophication in coastal river basins, the eutrophication status, physicochemical properties, and bacterial parameters of overlying waters and sediments in the catchment (CA), estuarine (EA), and offshore (OA) areas in the Duliujian River Basin of Bohai Bay were investigated. The results showed that the eutrophication index (EI) of CA, EA, and OA were 62.71, 57.86, and 36.51, respectively. The EI was more sensitive to increases in P (slope = 3.887) than to increases in N (slope = 0.734) of the overlying water, indicating that P is the main factor driving eutrophication in the coastal river basin. However, a nonlinear relationship was found between P in sediments and overlying waters, suggesting that bacterial mediation may occur during P release. As speculated in this study, P in the overlying water increased more quickly than N with increasing bacterial diversity and metabolic abundance, indicating that the allometric release of N and P mediated by bacteria increases the risk of eutrophication. Redundancy analysis showed that organic matter and total N in sediment have positive effects on bacterial communities, which explains 21.8% and 23.7% variation in bacterial diversity, and explains 31.3% and 7.1% variation in bacterial metabolism. This also suggests that the accumulation of N in the sediment promotes the release of P and further aggravates the eutrophication of water. Therefore, simultaneous control of N and P is necessary to control water eutrophication in coastal river basins.

What Is the Appropriate Acupuncture Treatment Schedule for Chronic Pain? Review and Analysis of Randomized Controlled Trials.

BACKGROUND: Acupuncture is widely used for the treatment of chronic pain. Different protocols of acupuncture practice exist and lack agreement on the optimal schedule of acupuncture treatment. OBJECTIVE: To review the appropriate acupuncture treatment schedule for chronic pain. METHODS: Embase, Pubmed, Cochrane Central Register of Controlled Trials, and reference lists were searched from 2009 to 2018 to identify randomized controlled trials of acupuncture for chronic pain conditions. We collected factors of treatment schedule (D, duration of each treatment session; N, number of treatment sessions; T, total duration of treatment in weeks) from each of the trials, and the linear regression analysis with real pain relief rate (both treatment and follow-up) was performed. Furthermore, we recommend the concept of "DOSE" and frequency (F) to evaluate the dose and frequency effect of acupuncture. RESULTS: Twenty-four trials with a total number of 3461 patients met the inclusion criteria. Of these, data from 23 studies were available for analysis. Firstly, the results showed that follow-up pain relief rate was decreased slightly with the increase of the duration of each session and DOSE (r=-0.3414 and r=-0.3246, respectively), but those two factors had no correlation with the pain relief rate after treatment. Secondly, it showed that either lower frequency with 2 sessions/week and higher frequency greater than 2 sessions/week or DOSE of 30 mins/week can achieve higher pain relief rate after treatment. Thirdly, we found the rate of pain relief remained at a high level greater than 20% up to 18 weeks after the treatment, and then it dropped sharply below 10% with the follow-up extended. A positive relationship was found between study score and pain relief both in treatment and follow-up (r=0.4654 and r=0.3046, respectively). CONCLUSIONS: The effect of acupuncture varies greatly with the different schedules of acupuncture, so it is necessary to review and choose the appropriate schedule. Although the current work is based on a limited number of trials, the findings suggest that acupuncture has a dose and frequency effect presenting within a certain range, which would have considerable implications for the design and interpretation of clinical trials. More high-quality randomized controlled trials on acupuncture schedule research were needed for providing more definitive evidence.

Analysis of chemical composition and in vitro antidermatophyte activity of ethanol extracts of Dryopteris fragrans (L.) Schott.

ETHNOPHARMACOLOGICAL RELEVANCE: Dryopteris fragrans (L.) Schott is a deciduous perennial herb, which has been used traditionally for treatment of ringworm infections and others skin diseases in the north of China. AIM OF THE STUDY: To characterize the chemical composition, evaluate the antifungal activity and explore possible mechanisms about action of ethanol extracts of D. fragrans. MATERIALS AND METHODS: The chemical components in the ethanol extracts of D. fragrans were detected by high-performance liquid chromatography coupled with electrospray ionization and quadruple time-of-flight mass spectrometry (HPLC-ESI-QTOF-MS/MS). The minimal inhibitory concentrations (MIC) and minimal fungicidal concentrations (MFC) of the ethanol extracts of D. fragrans were determined by the Clinical and Laboratory Standards Institute (CLSI) M38-A2 method against 62 isolates of dermatophytes. The kinetics of fungal kill, synergy testing by checkerboard dilution and quantitation of sterol by ultra-performance liquid chromatography (UPLC) on Trichophyton rubrum and Trichophyton mentagrophytes were also investigated. RESULTS: Fourteen derivatives of phloroglucinol were identified in the ethanol extracts of D. fragrans. The MIC of the ethanol extracts of D. fragrans ranged from 0.059 to 3.780 mg/mL while MFC ranged from 0.118 to 3.780 mg/mL. The ethanol extracts of D. fragrans exerted fungicidal activity after 12 h of incubation against Trichophyton rubrum while it required 36 h of incubation against Trichophyton mentagrophytes at concentrations of 8 × MIC. In synergy testing, the interaction between miconazole (MCZ) and terbinafine (TBF) with the ethanol extracts of D. fragrans proved to be indifferent by testing fractional inhibitory concentration (FIC) values. Sterol in samples of fungal cells treated with the ethanol extracts of D. fragrans was significantly reduced. CONCLUSIONS: The ethanol extracts of D. fragrans had antifungal and fungicidal activity against dermatophytes and was likely a strain-dependent fungicidal agent. Interaction between drugs was indifferent on tested isolates. The inhibition of ergosterol biosynthesis was one of the antifungal mechanisms of the ethanol extracts of D. fragrans. These results showed that the ethanol extracts of D. fragrans could be explored for promising antifungal drugs. Dozens of phloroglucinol derivatives may contribute to high antifungal activity of the ethanol extracts of D. fragrans.

The effects of mycotoxins and selenium deficiency on tissue-engineered cartilage.

OBJECTIVE: To investigate the effects of 3 mycotoxins, deoxynivalenol (DON), nivalenol (NIV) and T-2 toxin, in the presence and absence of selenium (Se) on the metabolism of tissue-engineered cartilage to mimic conditions found in Kashin-Beck disease (KBD) environments. MATERIALS AND METHODS: Chondrocytes were seeded onto bone matrix gelatin (BMG) to construct engineered cartilage. The 3 toxins were added to the culture media for 3 weeks followed by immunhistochemical analyses of collagens type II and X, aggrecan, matrix metalloproteinases 1 and 3 (MMP-1 and MMP-3), MMP inhibitors 1 and 3 (TIMP-1 and TIMP-3) and α(2) macroglobulin (α2M). RESULTS: Type II collagen was decreased while type X collagen was increased in response to DON, NIV and T-2 toxin. Aggrecan was reduced by all 3 mycotoxins. Compared with the control, the 3 toxins decreased the expression of α2M, TIMP-1 and TIMP-3, and increased the expression of MMP-1 and MMP-3. Se could partially inhibit the effects of DON, NIV and T-2 toxins. CONCLUSION: Under the low Se condition, the 3 mycotoxins produced procatabolic changes in cartilage resulting in the loss of aggrecan and type II collagen and promoted a hypertrophic phenotype of chondrocytes characterized by increasing type-X-collagen expression, enhancing the expression of MMPs, while weakening the TIMPs. Se could partially block the effects mentioned above. These results support the hypothesis that the combination of mycotoxin stress and Se deficiency would be the causative factors for KBD.

Effects of T-2 toxin and selenium on chondrocyte expression of matrix metalloproteinases (MMP-1, MMP-13), α2-macroglobulin (α2M) and TIMPs.

T-2 toxin is regarded as an important etiological factor of Kashin-Beck disease, and supplementation of selenium-salt partly prevents Kashin-Beck disease. The present study investigated the effects of T-2 toxin on the degradation of type II collagen in human chondrocytes in vitro. Human chondrocytes were isolated and cultured on bone matrix gelatin to form an artificial cartilage model in vitro with or without T-2 toxin and selenium. Immunohistochemistry analyses showed that T-2 toxin decreased type II collagen staining and selenium appeared to prevent the decrease in type II collagen induced by T-2 toxin in engineered cartilage. Then, Western blot and RT-PCR analyses showed that an increase in MMP-13 and MMP-1 expressions, and a decrease in the expression of the general endoproteinase inhibitor (α(2)M) were induced by T-2 toxin. Gelatin reverse zymography showed that TIMP-1 and TIMP-2 levels were decreased in a dose-dependent manner after exposure of T-2 toxin. Selenium had a protective role by increasing the level of type II collagen protein through down-regulation of MMP-13 protein and mRNA expression and up-regulation of TIMP-1 and TIMP-2 expressions. These data suggest T-2 toxin induces cartilage matrix degradation by the up-regulation of MMP-13 and TIMP-1, and down-regulation of TIMP-2 and α(2)M expressions.