Diverse CMT2 neuropathies are linked to aberrant G3BP interactions in stress granules.

Complex diseases often involve the interplay between genetic and environmental factors. Charcot-Marie-Tooth type 2 neuropathies (CMT2) are a group of genetically heterogeneous disorders, in which similar peripheral neuropathology is inexplicably caused by various mutated genes. Their possible molecular links remain elusive. Here, we found that upon environmental stress, many CMT2-causing mutant proteins adopt similar properties by entering stress granules (SGs), where they aberrantly interact with G3BP and integrate into SG pathways. For example, glycyl-tRNA synthetase (GlyRS) is translocated from the cytoplasm into SGs upon stress, where the mutant GlyRS perturbs the G3BP-centric SG network by aberrantly binding to G3BP. This disrupts SG-mediated stress responses, leading to increased stress vulnerability in motoneurons. Disrupting this aberrant interaction rescues SG abnormalities and alleviates motor deficits in CMT2D mice. These findings reveal a stress-dependent molecular link across diverse CMT2 mutants and provide a conceptual framework for understanding genetic heterogeneity in light of environmental stress.

Graphdiyne oxide nanosheets display selective anti-leukemia efficacy against DNMT3A-mutant AML cells.

DNA methyltransferase 3 A (DNMT3A) is the most frequently mutated gene in acute myeloid leukemia (AML). Although chemotherapy agents have improved outcomes for DNMT3A-mutant AML patients, there is still no targeted therapy highlighting the need for further study of how DNMT3A mutations affect AML phenotype. Here, we demonstrate that cell adhesion-related genes are predominantly enriched in DNMT3A-mutant AML cells and identify that graphdiyne oxide (GDYO) display an anti-leukemia effect specifically against these mutated cells. Mechanistically, GDYO directly interacts with integrin ß2 (ITGB2) and c-type mannose receptor (MRC2), which facilitate the attachment and cellular uptake of GDYO. Furthermore, GDYO binds to actin and prevents actin polymerization, thus disrupting the actin cytoskeleton and eventually leading to cell apoptosis. Finally, we validate the in vivo safety and therapeutic potential of GDYO against DNMT3A-mutant AML cells. Collectively, these findings demonstrate that GDYO is an efficient and specific drug candidate against DNMT3A-mutant AML.

tBHQ attenuates podocyte injury in diabetic nephropathy by inhibiting NADPH oxidase-derived ROS generation via the Nrf2/HO-1 signalling pathway.

Aims: Oxidative stress plays a crucial role in podocyte injury in diabetic nephropathy (DN). tert-Butylhydroquinone (tBHQ) is an activator of Nrf2 that exerts protective effects in diabetic mice, but the underlying mechanism of tBHQ in the podocytes of DN is not fully understood. Materials and methods: A high glucose (HG)-induced HK2 cell model and streptozotocin-induced rat model of DN were established and treated with tBHQ or apocynin. The expression levels of Nrf2, HO-1, NOX2 and NOX4 were determined by Western blot or immunohistochemical staining. The level of oxidative stress in podocytes or kidney tissues was assessed using DCFH-DA or dihydroethidium (DHE) staining. Cell injury was assessed by F-actin staining and flow cytometry analysis. Key findings: We showed that HG treatment increased the expressions of NOX2 and NOX4 and enhanced ROS production in podocytes. Inhibition of NADPH oxidase activity by apocynin dramatically attenuated HG-induced ROS production and further alleviated cell injury and apoptosis in podocytes. Moreover, we found that HG inhibited the Nrf2/HO-1 signalling pathway in podocytes; however, tBHQ treatment significantly activated the Nrf2 signalling pathway, inhibited NADPH oxidase activity, and attenuated ROS production and cell injury in HG-treated podocytes. Furthermore, we observed that tBHQ treatment partially attenuated renal injury, activated the Nrf2 signalling pathway, inhibited NADPH oxidase activity and reduced ROS generation in the kidneys of STZ-induced diabetic rats. Significance: These results suggest that tBHQ exerts a protective role in hyperglycaemia-induced podocyte injury, and that the potential protective mechanism of tBHQ involves inhibiting NADPH oxidase-derived ROS generation by activating the Nrf2/HO-1 signalling pathway.

Antileukemic effect of caffeic acid 3,4-dihydroxyphenetyl ester. Evidences for its mechanisms of action.

BACKGROUND: Caffeic acid 3,4-dihydroxyphenethyl ester (CADPE) is a natural polyphenolic ester isolated as a minor component from a water extract of the Chinese medicine Zhongjiefeng [Sarcandra glabra (Thunb.) Nakai (Chloranthaceae)] and has previously shown to have activity against solid tumors through the modulation of multiple targets or signal pathways. However, the activity and potential mechanism of CADPE against leukemia cells have not yet been characterized. PURPOSE: To investigate whether and how CADPE kills leukemia cells. METHOD: (1) The activity of CADPE inhibiting the growth of different leukemia cell lines was evaluated by MTT assay; (2) Cell cycle arrest and apoptosis induced by CADPE were determined by flow cytometry with FlowJo software for quantification; (3) The protein levels were analyzed by Western blot and ubiquitin-binding c-Myc was acquired by co-immunoprecipitation. RESULTS: CADPE exerted potent activity against different leukemia cell lines with low toxicity in normal cells. In terms of mechanism of action, CADPE promoted ubiquitin-proteasome-dependent degradation of c-Myc through activating glycogen synthase kinase-3ß (GSK3ß) and downregulating deubiquitinating enzyme USP28 to trigger the interaction of c-Myc with ubiquitin ligase Fbw7, resulting in the downregulation of cell cycle regulators and anti-apoptotic proteins and consequently, cell cycle arrest and cell apoptosis. CONCLUSION: CADPE is a novel c-Myc inhibitor with high activity and a unique mechanism for killing leukemia cells.

A small natural molecule CADPE kills residual colorectal cancer cells by inhibiting key transcription factors and translation initiation factors.

Residual disease is the major cause for colorectal cancer (CRC) relapse. Herein, we explore whether and how a natural molecule CADPE killed heterogenic populations in a panel of CRC cell lines with KRAS/BRAF mutations that are natively resistant to EGFR- or VEGFR-targeted therapy, without sparing persistent cells, a reservoir of the disease relapse. Results showed that CADPE killed the tumor bulk and residual cells in the panel of CRC cell lines, rapidly inactivated c-Myc, STAT3, and NF-κB, and then decreased the protein levels of key signaling molecules for CRC, such as ß-catenin, Notch1, and the nodes of mTOR pathways; eukaryotic translation initiation factors (eIF4F); anti-apoptotic proteins (Bcl-xl, Mcl-1, and survivin); and stemness-supporting molecules (CD133, Bim-1, and VEGF). In terms of mechanism of action, concurrent downregulation of Mcl-1, Bcl-xl, and survivin was necessary for CADPE to kill CRC bulk cells, while additional depletion of CD133 and VEGF proteins was required for killing the residual CRC cells. Moreover, the disabled c-Myc, STAT3, NF-κB, and eIF4F were associated with the broadly decreased levels of anti-apoptosis proteins and pro-stemness proteins. Consistently, CADPE suppressed CRC tumor growth associated with robust apoptosis and depleted levels of c-Myc, STAT3, NF-κB, eIF4F, anti-apoptotic proteins, and pro-stemness proteins. Our findings showed the promise of CADPE for treating CRC and suggested a rational polytherapy that disables c-Myc, STAT3, NF-κB, and eIF4F for killing CRC residual disease.

Novel Bioactive Penicipyrroether A and Pyrrospirone J from the Marine-Derived Penicillium sp. ZZ380.

The marine-sourced fungus Penicillium sp. ZZ380 was previously reported to have the ability to produce a series of new pyrrospirone alkaloids. Further investigation on this strain resulted in the isolation and identification of novel penicipyrroether A and pyrrospirone J. Each of them represents the first example of its structural type, with a unique 6/5/6/5 polycyclic fusion that is different from the 6/5/6/6 fused ring system for the reported pyrrospirones. Their structures were elucidated by extensive nuclear magnetic resonance (NMR) and high resolution electrospray ionization mass spectroscopy (HRESIMS) spectroscopic analyses, electronic circular dichroism (ECD) and 13C NMR calculations and X-ray single crystal diffraction. Penicipyrroether A showed potent antiproliferative activity against human glioma U87MG and U251 cells with half maximal inhibitory concentration (IC50) values of 1.64-5.50 µM and antibacterial inhibitory activity with minimum inhibitory concentration (MIC) values of 1.7 µg/mL against methicillin-resistant Staphylococcus aureus and 3.0 µg/mL against Escherichia coli.

An integrative model of internalized stigma and recovery-related outcomes among people diagnosed with schizophrenia in rural China.

PURPOSE: Internalized stigma, an adverse psychological process, severely impedes the lives of people diagnosed with schizophrenia and restricts them from social integration and recovery. The aim of this study was to empirically evaluate an integrative model of relationship between internalized stigma and patients' recovery-related outcomes among people diagnosed with schizophrenia in a rural Chinese community. METHOD: A total of 232 people diagnosed with schizophrenia in Xinjin, Chengdu, participated in this study and completed measures of internalized stigma, social interaction, perceived social support, social functioning, and symptoms. The internalized stigma of mental illness scale (ISMI) was used to measure the internalized stigma. Path analysis was used to test the association between internalized stigma and recovery-related outcomes. RESULTS: There were no significant differences in mean scores of ISMI by gender, age (18-64 years and ≥ 65 years), education, marital status, or economic capacity. Internalized stigma was negatively associated with perceived social support and social interaction. Furthermore, higher level of internalized stigma was associated with impaired social functioning, and a lower level of social functioning was significantly associated with more severe symptoms. CONCLUSION: Internalized stigma is associated with poor social interaction and weakened perceived social support in people diagnosed with schizophrenia, and is linked negatively to outcomes in their recovery. It is essential to tailor interventions related to reducing internalized stigma within a Chinese context and evaluate the effectiveness of anti-stigma intervention on recovery for people diagnosed with schizophrenia.

[Herb-drug enzyme-mediated interactions and the associated experimental methods: a review].

OBJECTIVE: To review the interactions between herbs and widely used drugs and summarize the associated experimental methods. METHODS: Definite herb-drug interactions were obtained by searching PubMed, other large overseas databases and summarizing new researches from China. We summarize some methods to assess the interaction between herbs and drugs involving microsomal, cell culture and animal experiments, and clinical trials, classifying this method as single ingredient herbs, crude herb extracts, and herbal formulae. RESULTS: Many herbs interact with drugs through a complex cytochrome P450 and/or P-glycoprotein mechanism. Herb-induced enzyme inhibition and/or induction may result in enhanced and/or decreased plasma, tissue, urine and bile drug concentrations, leading to a change in a drug's pharmacokinetic parameters and resulting in the improper treatment of patients and potentially severe side effects. Use of an appropriate method for comprehensively assessing herb-drug interactions can minimize clinical risks. Different methods were used by researchers to assess the pharmacological changes of drugs in vivo and in vitro and the mechanisms of the interactions from microsomal, cell culture and animal experiments, and clinical trials are discussed in this review. CONCLUSION: Co-medication with herbs can result in changes in pharmacological effects of many drugs. This review describes the assessment of single-ingredient herbs, crude herb extracts, and herbal formulae. When choosing a research method to investigate herb-drug interactions, the properties of the drugs and herbs should be considered.

Identification of the related substances in ampicillin capsule by rapid resolution liquid chromatography coupled with electrospray ionization tandem mass spectrometry.

Rapid Resolution Liquid Chromatography coupled with Electrospray Ionization Tandem Mass Spectrometry (RRLC-ESI-MS(n)) was used to separate and identify related substances in ampicillin capsule. The fragmentation behaviors of related substances were used to identify their chemical structures. Finally, a total of 13 related substances in ampicillin capsule were identified, including four identified components for the first time and three groups of isomers on the basis of the exact mass, fragmentation behaviors, retention time, and chemical structures in the literature. This study avoided time-consuming and complex chemosynthesis of related substances of ampicillin and the results could be useful for the quality control of ampicillin capsule to guarantee its safety in clinic. In the meantime, it provided a good example for the rapid identification of chemical structures of related substances of drugs.

[Construction of biopharmaceutics classification system of Chinese materia medica].

Based on the characteristics of multicomponent of traditional Chinese medicine and drawing lessons from the concepts, methods and techniques of biopharmaceutics classification system (BCS) in chemical field, this study comes up with the science framework of biopharmaceutics classification system of Chinese materia medica (CMMBCS). Using the different comparison method of multicomponent level and the CMMBCS method of overall traditional Chinese medicine, the study constructs the method process while setting forth academic thoughts and analyzing theory. The basic role of this system is clear to reveal the interaction and the related absorption mechanism of multicomponent in traditional Chinese medicine. It also provides new ideas and methods for improving the quality of Chinese materia medica and the development of new drug research.

[Design and analyze mathematical algorithms of intestinal absorption and metabolism of multicomponent drug].

Evaluation of the permeability mainly focuses on intestinal absorption in biopharmaceutics classification system (BCS). It is more complicated that the absorption and metabolism under multicomponent environment in biopharmaceutics classification system of Chinese materia medica (CMMBCS) compared with single component environment, which needs suitable mathematical models to be described. Therefore, with full consideration of existing single component mathematical algorithm combining with the characteristics of intestinal absorption and metabolism, we explored and designed a new mathematical algorithm of intestinal absorption and metabolism of multicomponent drug. Then we put forward a new coefficient, P (influence), the relative change rate of the single component's intestinal absorption and metabolism under multicomponent environment compared with single component environment, which described the influences of intestinal absorption and metabolism of the component under multicomponent environment. Moreover, P (influence) highlights the distinctive characteristics of multicomponent drug's intestinal absorption and metabolism, and lays the foundation for the construction of CMMBCS.

[Application of multicomponent dissolution evaluation method of biopharmaceutics classification system of Chinese materia medica in gegen qinlian tablets].

The study is a paticular embodiment of Chinese patent medicine based on biopharmaceutics classification system of Chinese materia medica (CMMBCS) , focusing on assessment of synchronization issues of dissolution that may affect the timing of the multicomponent absorption. The accumulative dissolution percentages of nine components in Gengen Qinlian tablets in different dissolution solvents and times were determined by HPLC. The dissolution curve was drew and its similarity was evaluated by similarity factors (f2) and cluster method. Results in this experiment showed that the components that peak 7 and peak 8 (baicalin) represented had poor similarity with the reference peak 2 (puerarin). Their similarity factors were both 43 in water dissolution media and 31 and 45 in pH 7.4 dissolution media, respectively. Components that peaks represented had better similarity with the reference peak 2 (puerarin) in other medium. It illustrated that components that peak 3,4,5,6 (berberine) represented had fully synchronous dissolution characteristics with the reference peak 2 (puerarin), components peak 1 and 9 represented had nearly fully synchronous dissolution characteristics with the reference peak 2 (puerarin), while components that peak 7 and 8 (baicalin) represented had no synchronous dissolution characteristics with the reference peak 2 (puerarin).

[Effect of multicomponent environment on intestinal permeability of puerarin in biopharmaceutics classification system of Chinese materia medica].

The evaluation of permeability in biopharmaceutics classification system of Chinese materia medica (CMMBCS) requires multicomponent as a whole in order to conduct research, even in the study of a specific component, should also be put in the multicomponent environment. Based on this principle, the high content components in Gegen Qinlian decoction were used as multicomponent environmental impact factors in the experiment, and the relevant parameters of intestinal permeability about puerarin were measured with using in situ single-pass intestinal perfusion model, to investigate and evaluate the intestinal permeability of puerarin with other high content components. The experimental results showed that different proportions of baicalin, glycyrrhizic acid and berberine had certain influence on intestinal permeability of puerarin, and glycyrrhizic acid could significantly inhibit the intestinal absorption of puerarin, moreover, high concentration of berberine could promote the absorption of puerarin. The research results indicated that the important research ideas of permeability evaluation in biopharmaceutics classification system of Chinese materia medica with fully considering the effects of other ingredients in multicomponent environment.

Ex vivo and in situ approaches used to study intestinal absorption.

Over the recent years, intestinal absorption has been recognized as a critical factor affecting the bioavailability of oral drugs. Intestinal absorption is affected by many factors including the physicochemical property of the drug, the absorption mechanisms, and the need for absorption enhancers. Ex vivo and in situ methods have been used extensively to evaluate the intestinal absorption of new drugs. Biological performance can be obtained rapidly and reliably using these techniques. However, these approaches have many inadequacies which need to be recognized so that appropriate adjustments can be made to the methodology. These shortcomings also need to be accounted for during the interpretation and application of the results in vivo situations. This review describes ex vivo and in situ models of drug absorption, and compares their relative advantages and drawbacks to assist researchers in selecting appropriate models for different drug and therapeutic situations.