Mechanisms of JinHong Formula on treating sepsis explored by randomized controlled trial combined with network pharmacology.

ETHNOPHARMACOLOGICAL RELEVANCE: JinHong Formula (JHF) was derived from the famous Rhubarb and Moutan Decoction which was prescribed for appendicitis. It was originally recorded in the classic of "Jingui Yaolve" written by Zhang Zhongjing. It is a kind of traditional Chinese medicine, widely used in the treatment of inflammation. However, the clinical effect of JHF for sepsis and its comprehensive mechanism in sepsis remained largely unknown. RESEARCH PURPOSE: The aim of our study was to evaluate the clinical effect of JHF in the treatment of sepsis, and to explore its mechanism from the perspective of network pharmacology. RESEARCH METHODS: The single-center randomized clinical trial was conducted to assess the effect of JHF in the treatment of sepsis. Additionally, we used the Chinese herbal medicine pharmacology database and analysis platform to identify the active components and therapeutic target of JHF. Numerous well-known disease target databases have been used to screen therapeutic target proteins for sepsis. Furthermore, we have established a Protein-Protein Interaction (PPI) network and carried out Gene Onotology/Kyoto Encyclopedia of Genes and Genomes (GO/KEGG) enrichment analysis. In order to conclude which active compounds from JHF may be responsible for signaling pathway, we performed network analysis. RESEARCH RESULTS: The study included 114 patients. By comparing participants with and without JHF, the results suggested that JHF significantly reduced all-cause mortality on 28 and 60 days after intervention, and improved Sequential Organ Failure Assessment (SOFA) on 7th day after intervention as well as. JHF had an effect of anti-inflammatories and antioxidants (SOD). By using network pharmacological analysis, we identified 72 active components and 426 target genes of JHF, and successfully constructed a "JHF-compound target-sepsis" network. 116 mentioned targets revealed by GO/KEGG enrichment analysis played a significant role in the inflammatory reaction and immunoregulation via interleukin-17 (IL-17) and tumor necrosis factor (TNF) signaling pathway. Moreover, the analysis of "pathway target-active component" revealed that Sennidin A, Rheidin A, Rheidin B, Rheidin C, (E)-4-Phenyl-3-Buten-2-One, Osmanthuside H, Esculetin, and Caffeicacid were responsible for IL-17, TNF signaling pathways. CONCLUSION: JHF contains potential active substance of anti-inflammatory and antioxidant. These active compounds may come into play through IL-17 and TNF signaling pathways. For sepsis, JHF may be a promising and effective treatment strategy.

Progress in infrared spectroscopy as an efficient tool for predicting protein secondary structure.

Infrared (IR) spectroscopy is a highly sensitive technique that provides complete information on chemical compositions. The IR spectra of proteins or peptides give rise to nine characteristic IR absorption bands. The amide I bands are the most prominent and sensitive vibrational bands and widely used to predict protein secondary structures. The interference of H2O absorbance is the greatest challenge for IR protein secondary structure prediction. Much effort has been made to reduce/eliminate the interference of H2O, simplify operation steps, and increase prediction accuracy. Progress in sampling and equipment has rendered the Fourier transform infrared (FTIR) technique suitable for determining the protein secondary structure in broader concentration ranges, greatly simplifying the operating steps. This review highlights the recent progress in sample preparation, data analysis, and equipment development of FTIR in A/T mode, with a focus on recent applications of FTIR spectroscopy in the prediction of protein secondary structure. This review also provides a brief introduction of the progress in ATR-FTIR for predicting protein secondary structure and discusses some combined IR methods, such as AFM-based IR spectroscopy, that are used to analyze protein structural dynamics and protein aggregation.

Highly efficient synergistic biocatalysis driven by stably loaded enzymes within hierarchically porous iron/cobalt metal-organic framework via biomimetic mineralization.

The integration of multimodal chemo-/bio-catalysis for efficient cascade reactions has long provided broad prospects in the field of biotechnology for ages. In this work, we describe the synthesis of a biomimetic multienzyme hybrid with hierarchically porous structure and outstanding catalytic activity via in situ encapsulation of natural enzymes in an iron-cobalt bimetallic metal-organic framework (Fe/Co-MOF, FCM). The combination of a single enzyme (glucose oxidase) or dual enzyme (ß-galactosidase and glucose oxidase) with FCM resulted in remarkable synergistic biocatalysis ability; in contrast to simple biocatalyst mixtures in solution, the prepared multienzyme hybrid resulted in 3.2-fold and 2.1-fold improvements in activity for tandem reactions, respectively. The reinforced cascade bioactivity of the multienzyme hybrid benefitted from the synergistic effect between iron/cobalt in the FCM nanozyme, the opened substrate channel between enzymes/nanozymes, and the beneficial effect provided by the hierarchical MOF pores. The enlarged pores not only provided adequate space for immobilized proteins to diffuse and reorientate in FCM with low surface energy, but also reduced the intrinsic mass transfer obstacle to increase the diffusional efficiency of reactants/intermediates. In addition, on account of the shielding effect provided by FCM, the multienzyme hybrid exhibited enhanced tolerance towards severe circumstances and excellent reusability and has been successfully applied in small molecule detection, such as glucose and lactose. The current study highlights the superiority of synergistic bioreactors integrated with the MOF nanozyme and natural enzymes, suggesting great potential for applications in sustainable biomimetic catalysis.

Therapeutic potential of endogenous hydrogen sulfide inhibition in breast cancer (Review).

Hydrogen sulfide (H2S), the third gas signal molecule, is associated with the modulation of various physiological and pathological processes. Recent studies have reevealed that endogenous H2S may promote proliferation, induce angiogenesis and inhibit apoptosis, thereby stimulating oncogenesis. Conversely, decreased endogenous H2S release suppresses growth of various tumors including breast cancer. This observation suggests an alternative tumor therapy strategy by inhibiting H2S­producing enzymes to reduce the release of endogenous H2S. Breast cancer is the most common type of cancer in women. Due to the lack of approved targeted therapy, its recurrence and metastasis still affect its clinical treatment. In recent years, significant progress has been made in the control of breast cancer by using inhibitors on H2S­producing enzymes. This review summarized the roles of endogenous H2S­producing enzymes in breast cancer and the effects of the enzyme inhibitors on anticancer and anti­metastasis, with the aim of providing new insights for the treatment of breast cancer.

Microfluidic Raman biochip detection of exosomes: a promising tool for prostate cancer diagnosis.

Tumor-derived exosomes, which contain RNA, DNA, and proteins, are a potentially rich non-invasive source of biomarkers. However, no efficient isolation or detection methods are yet available. Here, we developed a microfluidic Raman biochip designed to isolate and analyze exosomes in situ. Anti-CD63 magnetic nanoparticles were used to enrich exosomes through mixing channels of a staggered triangular pillar array. EpCAM-functionalized Raman-active polymeric nanomaterials (Raman beads) allow rapid analysis of exosome samples within 1 h, with a quantitative signal at 2230 cm-1. The limit of detection of this biochip approaches 1.6 × 102 particles per mL with 20 µL samples. The newly developed biochip assay was successfully applied in the determination of exosomes from clinical serum samples. Thus, this novel device may have potential as a clinical exosome analysis tool for prostate cancer.

Cystathionine­Î³­lyase promotes the metastasis of breast cancer via the VEGF signaling pathway.

The present study aimed to provide data to support the association between cystathionine­Î³­lyase (CSE) and breast cancer metastasis. Reverse transcription­quantitative polymerase chain reaction, immunohistochemistry and western blot analysis were used to detect the mRNA and protein expression levels of CSE in human breast cancer tissues and cells. MTS and 5­ethynyl­2'­deoxyuridine assays were used to assess cell viability and proliferation. Scratch wound and Transwell assays were conducted to determine cell migration and invasion. In addition, hydrogen sulfide determination was performed using the methylene blue method. The expression of CSE was upregulated in samples from patients with breast cancer that also exhibit lymph node metastasis, and in grade III and readily metastatic breast cancer cell lines. The proliferation, migration and invasion of breast cancer cells were examined in the present study, and tumor metastasis was observed in nude mice. The function of CSE in breast cancer metastasis depends on the vascular endothelial growth factor (VEGF) signaling pathway, a key mediator of angiogenesis that is crucial for the development and metastasis of tumors. CSE positively regulated the expression of VEGF and increased the levels of certain key proteins in the VEGF pathway, including the phosphoinositide (PI3K)/protein kinase B (AKT) pathway [PI3K, Akt and phosphorylated (p)Akt], focal adhesion kinase (FAK)­paxillin pathway (FAK and paxillin) and rat sarcoma (Ras)­mitogen­activated protein kinase pathway [Ras, rapidly accelerated fibrosarcoma, extracellular signal­regulated kinase (ERK)1/2 and pERK1/2]. Furthermore, the novel CSE inhibitor I157172 possessed antiproliferative and anti­metastatic activities in early MDA­MB­231 metastatic breast cancer cells via inhibition of the VEGF signaling pathway, which further confirmed the role of CSE in breast cancer metastasis. Overall, these data demonstrate for the first time, to the best of our knowledge, that the functions of CSE in breast cancer metastasis are associated with the VEGF signaling pathway.

I157172, a novel inhibitor of cystathionine γ-lyase, inhibits growth and migration of breast cancer cells via SIRT1-mediated deacetylation of STAT3.

Cystathionine γ-lyase (CSE) is highly expressed in breast cancer, and can promote breast cancer development and progression; therefore, inhibitors of CSE may be of great significance for the treatment of breast cancer. The present study identified the CSE inhibitor I157172 through virtual screening and confirmed its activity. Subsequently, the effects and mechanism of I157172 on breast cancer cells were investigated. MTS and 5-ethynyl-2'-deoxyuridine (EdU) assays were used to assess cell viability and proliferation. Scratch wound and Transwell assays were conducted to determine cell migration and invasion. In addition, H2S determination was performed using the methylene blue method, and western blotting was performed to detect protein expression. The results revealed that I157172 significantly inhibited the growth, proliferation and migration of MCF7 breast cancer cells in a dose-dependent manner. The results of further mechanistic studies demonstrated that CSE expression was negatively associated with sirtuin 1 (SIRT1) in human breast cancer tissues and cells, and CSE knockdown resulted in an increase in SIRT1 expression, and a decrease in acetylated (acetyl)-signal transducer and activator of transcription 3 (STAT3) and phosphorylated (p)-STAT3 levels in MCF7 cells. Furthermore, STAT3 downstream proteins B-cell lymphoma 2, p-protein kinase B, matrix metalloproteinase (MMP)-2 and MMP-9 were inhibited in CSE knockdown MCF7 cells. In addition, I157172 induced upregulation of SIRT1, and downregulation of acetyl-STAT3 and p-STAT3 (Tyr705), as well as inhibition of STAT3 downstream proteins. Taken together, I157172 inhibited the growth, proliferation and migration of breast cancer cells via upregulating SIRT1, which consequently mediated deacetylation of STAT3 and inactivation of the STAT3 pathway.

A label-free colorimetric isothermal cascade amplification for the detection of disease-related nucleic acids based on double-hairpin molecular beacon.

K-Ras mutations at codon 12 play an important role in an early step of carcinogenesis. Here, a label-free colorimetric isothermal cascade amplification for ultrasensitive and specific detection of K-Ras point mutation is developed based on a double-hairpin molecular beacon (DHMB). The biosensor consists of DHMB probe and a primer-incorporated polymerization template (PPT) designed partly complementary to DHMB. In the presence of polymerase, target DNA is designed to trigger strand displacement amplification (SDA) via promote the hybridization of PPT with DHMB and subsequently initiates cascade amplification process with the help of the nicking endonuclease. During the hybridization and enzymatic reaction, G-quadruplex/hemin DNAzymes are generated, catalyzing the oxidation of ABTS2- by H2O2 in the presence of hemin. Utilizing the proposed facile colorimetric scheme, the target DNA can be quantified down to 4 pM with the dynamic response range of 5 orders of magnitude, indicating the substantially improved detection capability. Even more strikingly, point mutation in K-ras gene can be readily observed by the naked eye without the need for the labeling or expensive equipment. Given the high-performance for K-Ras analysis, the enhanced signal transduction capability associated with double-hairpin structure of DHMB provides a novel rout to screen biomarkers, and the descripted colorimetric biosensor seems to hold great promise for diagnostic applications of genetic diseases.

[The effect of hydrogen on hemorrhagic shock induced acute lung injury in rats].

OBJECTIVE: To investigate the effect of hydrogen inhalation on acute lung injury after hemorrhagic shock in rats. METHODS: Twenty-four adult male Sprague-Dawley (SD) rats were equally randomized into three groups: sham operation group, model group and hydrogen-treatment group. Pressure-controlled hemorrhagic shock and resuscitation model was reproduced by blood-letting for 1 hour followed by fluid replacement for 2 hours. The rats in model group received a mixture of 50% oxygen-50% nitrogen during the process. The rats in hydrogen-treatment group received inhalation of a mixture of 2% hydrogen-48% nitrogen-50% oxygen 10 minutes before fluid replacement till the end of resuscitation. The arterial blood samples were collected for the measurement of arterial partial pressure of oxygen (PaO2) before exsanguination, 1 hour after shock, 1 hour and 2 hours after fluid replacement. Blood and lung tissues were collected at the end of experiment, and tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) levels in plasma, lung wet/dry weight ratio (W/D), malondialdehyde (MDA) content, superoxide dismutase (SOD) and myeloperoxidase (MPO) activity in the lung tissue were determined. The lung tissue was subjected to pathological examination. RESULTS: At the end of fluid replacement, compared with model group, hydrogen could significantly reduce pulmonary edema (lung W/D ratio: 4.72 ± 0.12 vs. 4.94 ± 0.14, P<0.05), inhibit oxidative stress (MDA: 0.55 ± 0.09 nmol/mg vs. 0.72 ± 0.08 nmol/mg, P<0.05), enhance antioxidant activity (SOD activity: 79.53 ± 14.33 U/mg vs. 59.55 ± 9.07 U/mg, P<0.05), reduce the release of pro-inflammatory cytokines (TNF-α: 55.58 ± 10.06 ng/L vs. 66.58 ± 5.17 ng/L; IL-6: 23.00 ± 2.77 ng/L vs. 27.09 ± 2.46 ng/L, both P<0.05) and inhibit neutrophil infiltration (MPO: 1.05 ± 0.18 U/g vs. 1.40 ± 0.14 U/g, P<0.05). It alleviated the damage to lung tissue, and then improved the lung function (PaO2: 146.3 ± 22.1 mm Hg vs. 123.6 ± 16.0 mm Hg, P<0.05). CONCLUSIONS: Hydrogen treatment can alleviate acute lung injury as a result of hemorrhagic shock and resuscitation.