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Background: In recent years, many naphthoquinone compounds with anticancer activity have been identified in Arnebiae Radix, and some of them have the potential to be developed into anticancer drugs. Objective: This article aimed to provide a comprehensive overview of the anticancer effects of naphthoquinone compounds through a detailed review of literature and Chinese patents, and discuss their potential to be developed as anticancer drugs for clinical application. Methods: Research papers were collected through the databases of PubMed, Cnki and SciDirect using keyword searches “naphthoquinone compounds” and “anticancer”. The keywords of “shikonin” and “shikonin derivatives” were also used in PubMed, Cnki and SciDirect databases to collect research articles. The Chinese patents were collected using the Cnki patent database. Results: Naphthoquinone compounds have been found to possess anti-cancer activity, and their modes of action are associated with inducing apoptosis, inhibiting cancer cell proliferation, promoting autophagy in cancer cells, anti-cancer angiogenesis and inhibition of cell adhesion, invasion and metastasis, inhibiting glycolysis and inhibiting DNA topoisomerase activity. Conclusion: Most of the naphthoquinone compounds show effective anti-cancer activity in vitro. The structure modification of naphthoquinone aims to develop anti-cancer drugs with high efficacy and low toxicity.

Background Accurate posterior column reduction remains a challenging and controversial topic in the management of complex pilon fractures (AO/OTA C3). We aim to report the outcomes of surgical treatment for 22 AO/OTA C3 pilon fracture cases between January 2015 and May 2017 and highlight some traps and tips. Methods Three patients underwent two-stage early plating on the posterior column through a posterolateral approach. The remaining 19 patients were treated with two-stage delayed plating on the posterior column: 11 patients were treated with a posterolateral approach, five patients with a modified posteromedial approach, and three patients with a single anterior approach. The reduction of the posterior column was evaluated according to the Burwell-Charnley’s radiographic criteria, and functional outcomes were assessed using the American Orthopedic Foot and Ankle Society (AOFAS) scores. Results Posterior column malreduction occurred in five cases, including in one case that was re-adjusted immediately and in another case that was re-adjusted during a two-staged delayed operation. According to Burwell-Charnley’s criteria, the satisfactory rate of fracture reduction was 81.8%. After 1 year, the mean AOFAS score was 81.9 (81.9 ± 9.9); the outcome was excellent in three (20.0%), good in nine (60.0%), and fair in three (20.0%). Excellent or good outcomes were noted in 12 patients (80.0%). Conclusions The combined anterior and posterior approach is suggested in the second stage of plating so that the posterior column fragments can be re-adjusted intraoperatively, if necessary. Following these procedures, satisfactory reduction and recovery of good ankle function can be anticipated.

Coeliac disease (CD), also known as gluten sensitive enteropathy, is an autoimmune intestinal disease induced by gluten in genetically susceptible individuals. Gluten is a common ingredient in daily diet and is one of the main environmental factors to induce coeliac disease. Adhering to gluten free diet (GFD) is an effective method for treating CD. Microbiota plays an extremely important role in maintaining human health, and diet is the main factor to regulate the composition and function of gut microbiota. Recent studies have shown that gluten metabolism is closely related to gastrointestinal tract (GIT) microbiota. With the increasing prevalence of coeliac disease, there is a need for alternative treatments to GFD. In this review, biological medication of gluten, relationship between gluten and gut microflora, effect of GFD on GIT microflora, and effect of probiotics on CD were reviewed. By analysing the research progress on relationship between gluten and gastrointestinal microbiome in coeliac disease, this review tried to explore the prospective and potential mechanism of microecological agents in treating coeliac disease.

In the process of ischemic stroke (IS), cellular macroautophagy/autophagy and apoptosis play a vital role in neuroprotection against it. Therefore, regulating their balance is a potential therapeutic strategy. It has been proved that hydroxysafflor yellow A (HSYA) has anti-inflammatory and antioxidant effects, which can both protect neurons. By exploring bioinformatics combined with network pharmacology, we found that HIF1A and CASP3, key factors regulating autophagy and apoptosis, may be important targets of HSYA for neuroprotection in an oxygen glucose deprivation and reperfusion (OGD/R) model. In this study, we explored a possible new mechanism of HSYA neuroprotection in the OGD/R model. The results showed that OGD/R increased the expression of HIF1A and CASP3 in SH-SY5Y cells and induced autophagy and apoptosis, while HSYA intervention further promoted the expression of HIF1A and inhibited the level of CASP3, accompanied by an increase in autophagy and a decrease in apoptosis in SH-SY5Y cells. The inhibition of HIF1A diminished the activation of autophagy induced with HSYA, while the inhibition of autophagy increased cell apoptosis and blocked the neuroprotective effect of HSYA, suggesting that the neuroprotective effect of HSYA should be mediated by activating the HIF1A/BNIP3 signaling pathway to induce autophagy. These results demonstrate that HSYA may be a promising agent for treating IS.

OBJECTIVE: To summarize the surgical learning curve and preliminary operative experience of dual-robotic navigated minimally invasive treatment on pelvic fractures by TiRobot and Artis Zeego. METHODS: Between July 2019 and February 2021, 90 patients with pelvic fractures were treated with dual-robotic navigated minimally invasive surgery by TiRobot and Artis Zeego. There were 64 males and 26 females, with an average age of 46.5 years (range, 13-78 years). Body mass index was 14.67-32.66 kg/m(2) (mean, 23.61 kg/m(2)). Causes of injuries included traffic accident in 43 cases, falling from height in 37 cases, low-energy injuries such as flat falls in 10 cases. The interval between injury and surgery was 1-36 days (mean, 7.3 days). According to the location of the implanted screws, the patients were divided into sacroiliac screw group (n=33), acetabular screw group (acetabulum anterior/posterior column, n=24), composite screws group (sacroiliac and acetabulum anterior/posterior column, n=33). According to the screw implantation time and accuracy, the surgical learning curve was plotted, and the differences in the relevant indicators between learning stage and skilled stage were compared. RESULTS: All 90 patients successfully completed the operation, the intraoperative bleeding volume was 5-200 mL (median, 20 mL). There was no vascular or nerve injury. All incisions healed by first intention. The screw implantation time ranged from 7.5 to 33.0 minutes (mean, 18.92 minutes), and the screw implantation accuracy ranged from 1.1 to 1.8 mm (mean, 1.56 mm). According to the learning curve, the practice stage of 3 groups was reached after 7, 10, and 11 cases, respectively. With the accumulation of surgical experience, the screw implantation time had a significant downward trend. Compared with the learning stage, the screw implantation time on skilled stage in 3 groups significantly shortened (P0.05). CONCLUSION: TiRobot and Artis Zeego assisted pelvic fracture surgery is safe and efficient, which helps the surgeon to quickly master the pelvic channel screw surgery, and the operation time is significantly shortened on the premise of ensuring the implantation accuracy.

Moran Huang,1,* Kai Ye,1,* Tu Hu,2,* Kexin Liu,3 Mengzhen You,3 Lei Wang,1 Hui Qin1 1Department of Orthopaedics, Shanghai Jiaotong University Affiliated Sixth People’s Hospital, Shanghai, People’s Republic of China; 2Department of Musculoskeletal Surgery, Fudan University Shanghai Cancer Center, Shanghai, People’s Republic of China; 3Department of General Practice, Shanghai Jiaotong University Affiliated Sixth People’s Hospital, Shanghai, People’s Republic of China*These authors contributed equally to this workCorrespondence: Hui Qin; Lei WangDepartment of Orthopaedics, Shanghai Jiaotong University Affiliated Sixth People’s Hospital, No. 600 Yishan Road, Shanghai, People’s Republic of ChinaTel +86-135-64143033; +86-189-30172609Fax +8621-6470-1361Email qh197826@163.com; wanglei2264@126.comPurpose: Despite the extensive development of antibacterial biomaterials, there are few reports on the effects of materials on the antibacterial ability of the immune system, and in particular of neutrophils. In this study, we observe differences between the in vivo and in vitro anti-infective efficacies of silver nanoparticles (AgNPs). The present study was designed to further explore the mechanism for this inconsistency using ex vivo models and in vitro experiments.Methods: AgNPs were synthesized using the polyol process and characterized by transmission electron microscopy and X-ray photoelectron spectroscopy. The antibacterial ability of AgNPs and neutrophils was tested by the spread-plate method. The infected air pouch model was prepared to detect the antimicrobial ability of AgNPs in vivo. Furthermore, blood-AgNPs-bacteria co-culture model and reactive oxygen species (ROS) measurement were used to evaluate the effect of AgNPs to neutrophil-mediated phagocytosis and ROS production.Results: The antibacterial experiments in vitro showed that AgNPs had superior antibacterial properties in cell compatible concentration. While, AgNPs had no significant antibacterial effect in vivo, and pathological section in AgNPs group indicated less neutrophil infiltration in inflammatory site than S. aureus group. Furthermore, AgNPs were found to reduce the phagocytosis of neutrophils and inhibit their ability to produce ROS and superoxide during ex vivo and in vitro experiments.Conclusion: This study selects AgNPs as the representative of inorganic nano-biomaterials and reveals the phenomenon and the mechanism underlying the significant AgNPs-induced inhibition of the antibacterial ability of neutrophils, and may have a certain enlightening effect on the development of biomaterials in the future. In the fabrication of antibacterial biomaterials, however, attention should be paid to both cell and immune system safety to make the antibacterial properties of the biomaterials and innate immune system complement each other and jointly promote the host’s ability to resist the invasion of pathogenic microorganisms.Keywords: antibacterial biomaterials, polymorphonuclear neutrophils, silver nanoparticles, immune system

Rivaroxaban, an oral anticoagulant, undergoes the metabolism mediated by human cytochrome P450 (CYP). The present study is to quantitatively analyze and compare the contributions of multiple CYPs in the metabolism of rivaroxaban to provide new information for medication safety. The metabolic stability of rivaroxaban in the presence of human liver microsomes (HLMs) and recombinant CYPs was systematically evaluated to estimate the participation of various CYP isoforms. Furthermore, the catalytic efficiency of CYP isoforms was compared via metabolic kinetic studies of rivaroxaban with recombinant CYP isoenzymes, as well as via CYP-specific inhibitory studies. Additionally, docking simulations were used to illustrate molecular interactions. Multiple CYP isoforms were involved in the hydroxylation of rivaroxaban, with decreasing catalytic rates as follows: CYP2J2 > 3A4 > 2D6 > 4F3 > 1A1 > 3A5 > 3A7 > 2A6 > 2E1 > 2C9 > 2C19. Among the CYPs, 2J2, 3A4, 2D6, and 4F3 were the four major isoforms responsible for rivaroxaban metabolism. Notably, the intrinsic clearance of rivaroxaban catalyzed by CYP2J2 was nearly 39-, 64-, and 100-fold that catalyzed by CYP3A4, 2D6, and 4F3, respectively. In addition, rivaroxaban hydroxylation was inhibited by 41.1% in the presence of the CYP2J2-specific inhibitor danazol, which was comparable to the inhibition rate of 43.3% by the CYP3A-specific inhibitor ketoconazole in mixed HLMs. Furthermore, molecular simulations showed that rivaroxaban is principally bound to CYP2J2 by π-alkyl bonds, carbon-hydrogen bonds, and alkyl interactions. CYP2J2 dominated the hydroxylation of rivaroxaban, which may provide new insight into clinical drug interactions involving rivaroxaban.

Mitochondria play a central role in biological oxidation that inevitably generates reactive oxygen species (ROS) as by-products. Maintenance of mitochondrial redox balance status requires NADPH, which is primarily generated by the mitochondrial matrix protein isocitrate dehydrogenase 2 (IDH2). The activity of IDH2 is regulated by post-translational modifications (PTMs). In this study, we found IDH2 is modified by small ubiquitin-like modifier 1 (SUMO1) at lysine 45. SUMO specific protease 1 (SENP1) is responsible for deSUMOylation of IDH2. SUMOylation of IDH2 is induced by oxidants and enhances the antioxidant activity of IDH2 to protect cells against oxidative stress. Mutation of the SUMOylation site impairs the enzymatic activity of IDH2 and hence decreases levels of α-ketoglutarate (α-KG), NADPH and GSH. Cells with SUMOylation deficient IDH2 suffer more apoptosis than that with wild type IDH2 under oxidative stress. These results indicate that SUMOylation is an important way to regulate IDH2 activity to maintain mitochondrial redox balance.

Imipenem (Imp) is a widely used broad-spectrum antibiotic. However, renal adverse effects limit its clinical application. We previously reported that organic anion transporters (OATs) facilitated the renal transport of Imp and contributed its nephrotoxicity. Natural flavonoids exhibited renal protective effect. Here, we aimed to develop potent OAT inhibitors from traditional Chinese medicines (TCMs) and to evaluate its protective effect against Imp-induced nephrotoxicity. Among 50 TCMs, Tribuli Fructus, Platycladi Cacumen, and Lycopi Herba exhibited potent inhibition on OAT1/3. After screening their main components, Apigenin strongly inhibited Imp uptake by OAT1/3-HEK293 cells with IC50 values of 1.98 ± 0.36 μM (OAT1) and 2.29 ± 0.88 μM (OAT3). Moreover, Imp exhibited OAT1/3-dependent cytotoxicity, which was alleviated by Apigenin. Furthermore, Apigenin ameliorated Imp-induced nephrotoxicity in rabbits, and reduced the renal secretion of Imp. Apigenin inhibited intracellular accumulation of Imp and sequentially decreased the nephrocyte toxicity in rabbit primary proximal tubule cells (rPTCs). Apigenin, a flavone widely distributed in TCMs, was a potent OAT1/3 inhibitor. Through OAT inhibition, at least in part, Apigenin decreased the renal exposure of Imp and consequently protected against the nephrotoxicity of Imp. Apigenin can be used as a promising agent to reduce the renal adverse reaction of Imp in clinic.

Background and purpose Diclofenac is a widely used nonsteroidal anti-inflammatory drug. However, adverse effects in the kidney limit its clinical application. The present study was aimed to evaluate the potential effect of cilastatin on diclofenac-induced acute kidney injury and to clarify the potential roles of renal organic anion transporters (OATs) in the drug-drug interaction between cilastatin and diclofenac. Experimental approach The effect of cilastatin was evaluated in diclofenac-induced acute kidney injury in mice. Human OAT1/3-transfected HEK293 cells and renal primary proximal tubule cells (RPTCs) were used to investigate OAT1/3-mediated transport and the cytotoxicity of diclofenac. Key results Cilastatin treatment decreased the pathological changes, renal dysfunction and elevated renal levels of oxidation products, cytokine production and apoptosis induced by diclofenac in mice. Moreover, cilastatin increased the plasma concentration and decreased the renal distribution of diclofenac and its glucuronide metabolite, diclofenac acyl glucuronide (DLF-AG). Similarly, cilastatin inhibited cytotoxicity and mitochondrial damage in RPTCs but did not change the intracellular accumulation of diclofenac. DLF-AG but not diclofenac exhibited OAT-dependent cytotoxicity and was identified as an OAT1/3 substrate. Cilastatin inhibited the intracellular accumulation and decreased the cytotoxicity of DLF-AG in RPTCs. Conclusion and implications Cilastatin alleviated diclofenac-induced acute kidney injury in mice by restoring the redox balance, suppressing inflammation, and reducing apoptosis. Cilastatin inhibited OATs and decreased the renal distribution of diclofenac and DLF-AG, which further ameliorated the diclofenac-induced nephrotoxicity in mice. Cilastatin can be potentially used in the clinic as a therapeutic agent to alleviate the adverse renal reaction to diclofenac.

Food allergens are closely related to their gastrointestinal digestion fate, but the changes in food allergens during digestion and related mechanisms are quite complicated. This review presents in detail digestion models for predicting allergenicity, the fates of food allergens in oral, gastric and duodenal digestion, and the applications of digestomics in mapping IgE-binding epitopes of digestion-resistant peptides. Moreover, this review highlights the structure-activity relationships of food allergens during gastrointestinal digestion. Digestion-labile allergens may share common structural characteristics, such as high flexibility, rendering them easier to be hydrolyzed into small fragments with decreased or eliminated allergenicity. In contrast, the presence of disulfide bonds, tightly wound α-helical structures, or hydrophobic domains in food allergens helps them resist gastrointestinal digestion, stabilizing IgE-binding epitopes, thus maintaining their sensitization. In rare cases, digestion leads to increased allergenicity due to exposure of new epitopes. Finally, the action of the food matrix and processing on the digestion and allergenicity of food allergens as well as the underlying mechanisms was overviewed. The food matrix can directly act on the allergen by forming complexes or new epitopes to affect its gastrointestinal digestibility and thereby alter its allergenicity or indirectly affect the allergenicity by competing for enzymatic cleavage or influencing gastrointestinal pH and microbial flora. Several processing techniques attenuate the allergenicity of food proteins by altering their conformation to improve susceptibility to degradation by digestive enzymes. Given the complexity of food components, the food itself rather than a single allergen should be used to obtain more accurate data for allergenicity assessment. PRACTICAL APPLICATION: The review article will help to understand the relationship between food protein digestion and allergenicity, and may provide fundamental information for evaluating and reducing the allergenicity of food proteins.

Asthma is accompanied by inflammatory progression. Macrophages are a major type of cells to response inflammation caused by different type of factors by polarized into specific phenotypes. Luteolin and glycyrrhizic acid exert protect role in asthma; however, their role in THP-1 derived macrophages polarization whether through regulating the expression of hsa_circ_0001326 is still unknown. The effect of luteolin and glycyrrhizic acid on THP-1 derived macrophages polarization were evaluated using qRT-PCR, Western blotting, and ELISA assay. The function of hsa_circ_0001326 on macrophages polarization in luteolin treated THP-1 derived macrophages were assessed after silence of hsa_circ_0001326. And the expression of its' potential downstream gene, including hsa-miR-136-5p and ubiquitin-specific protease 4 (USP4), were detected using qRT-PCR and Western blot analysis. Furthermore, the potential mechanism of hsa_circ_0001326 were validated using rescue experiment. Results showed that luteolin promoted M2 polarization and inhibited M1 polarization in THP-1 induced macrophages, but glycyrrhizic acid had no these effects. Hsa_circ_0001326 expression was upregulated in luteolin treat THP-1 derived macrophages. Silence of hsa_circ_0001326 reversed the function of luteolin on macrophages polarization. In addition, hsa_circ_0001326 attenuated the inhibition effect of luteolin on hsa-miR-136-5p expression, and the promotion effect on USP4 expression. Furthermore, hsa-miR-136-5p inhibitor reversed the effect of hsa_circ_0001326 on macrophages polarization and the USP4 expression. Taken together, luteolin activates M2 macrophages and suppresses M1 macrophages by upregulation of hsa_circ_0001326. Further mechanism maybe by regulating hsa_circ_0001326 downstream gene expression, including hsa-miR-136-5p and USP4, in THP-1 derived macrophages. These findings provide a new insight for macrophage polarization under stimulation of luteolin.

Osteoarthritis (OA) is the most common joint disease and is the leading cause of chronic disability among older people. Chondrocyte death and extracellular matrix (ECM) degradation was involved in OA pathogenesis. Ferroptosis was an iron-dependent cell death associated with peroxidation of lipids. Here, we proved that ferroptosis exists in OA and identified glutathione peroxidase 4 (GPX4) as an important regulator of OA.Ferroptosis-related alterations were analyzed in human OA and undamaged cartilage. Expression of GPX4 was examined in 55 paired human OA samples. Ferrostatin-1 (Fer-1) and Deferoxamine (DFO) were used to treat OA, in vitro and in vivo. Alterations of GPX4-mediated signaling pathway were identified by RNA-seq analysis. AAV-Gpx4-shRNA were used to downregulate GPX4 expression in vivo.Transcriptomic, biochemical, and microscopical analyses indicated that ferroptosis was closely associated with OA. Expression of GPX4 in the OA cartilage from 55 OA patients were significantly lower than undamaged cartilage. Fer-1 and DFO could protect OA in a necroptosis-independent manner, suggesting that ferroptosis exists in OA prog. Importantly, GPX4 downregulation could increase the sensitivity of chondrocytes to oxidative stress and aggravate ECM degradation through the MAPK/NFκB pathway. Furthermore, downregulation of GPX4 expression by AAV-Gpx4 shRNA aggravated OA in vivo.Ferroptosis contributes to OA pathogenesis and GPX4 was the intersection of two mechanisms in regulating OA progression: ferroptosis and ECM degradation.This work was supported by the Projects of International Cooperation and Exchanges of National Natural Science Funding of China (Grant no. 81820108020), the National Key Research and Development Project of China (Grant no. 2018YFC1106300) and the Basic Science Program of Shanghai Jiao Tong University Affiliated Sixth People's Hospital (Grant no. ynms202102).

Background: Cytochrome P450 1B1(CYP1B1) is an extrahepatic P450 isoenzyme that can participate in processes of undermining the effectiveness and safety of anti-cancer therapy. Ginsenosides are the main active ingredients in ginseng, which possesses rich pharmacological activities, including anti-cancer activity and organ protection. However, the effect of ginsenosides on the activity of CYP1B1 remains unclear. Objective: The present study aimed to investigate the inhibitory effect of ginsenosides on CYP1B1 and reveal the structure-inhibitory activity relationship. Methods: Firstly, recombinant CYP1B1 and EROD reactions were used to evaluate the inhibitory effect of ginsenosides. Secondly, molecular docking was used to simulate the interactions between ginsenosides and CYP1B1. Finally, the structure-inhibitory activity relationship was analyzed. Results: The ginsenosides, Rb2, Rd, and Rg3, significantly inhibited CYP1B1; the ginsenoside Rd showed the strongest inhibition effect, with a Ki value of 47.37 μM in non-competitive mode. Notably, ginsenoside Rd formed hydrogen bonds with two key amino acid residues of CYP1B1, and one bond was between the glycosyl in position 20 and ALA330, which also made ginsenoside Rd close to the heme iron of CYP1B1. In contrast, ginsenosides, Rb2 and Rg3, which showed weaker inhibition, interacted with only one CYP1B1 residue by the hydrogen bond, which was far away from the heme iron. Finally, the structure-inhibitory activity relationship analysis demonstrated that the number of glycosyls in position 20 and the type of sapogenins in the ginsenoside structure are the key factors determining inhibitory activity. Meanwhile, ALA330 was a vital amino acid in the potent inhibition of CYP1B1 by ginsenosides. Conclusion: A structure-dependent inhibitory effect on CYP1B1 was revealed for ginsenosides, among which ginsenoside Rd showed the strongest inhibition due to its mono-glycosyl in position 20 of the ginsenoside parent structure. These findings would provide evidence for the synthesis of novel CYP1B1 inhibitors to augment the anti-cancer therapeutic effect.

Autophagy plays a key role in ischemic stroke, but its mechanism remains to be elucidated. In order to explore the effect of autophagy on ischemic stroke, bibliometric analysis and view tools are used to identify the directions of the global research trends and construct full view of the autophagy in ischemic stroke from 2006 to 2022. The research hotspots of autophagy related to ischemic stroke are visually analyzed and generated various visual maps to display publications, authors, sources, countries, organizations, and keywords. By bibliometric analysis, it can be seen that the investigations of autophagy in ischemic stroke is focused on both brain injury and neuroprotection. The impact of a variety of inflammatory factors and signaling pathways on autophagy following an ischemic stroke is also studied. Autophagy plays an important role in all phases of ischemic stroke. It is of great significance to guide the development of treatment plans for ischemic stroke.

Drug resistance is a critical obstacle to effective treatment in patients with chronic myeloid leukemia. To understand the underlying resistance mechanisms in response to imatinib mesylate (IMA) and adriamycin (ADR), the parental K562 cells were treated with low doses of IMA or ADR for 2 months to generate derivative cells with mild, intermediate, and severe resistance to the drugs as defined by their increasing resistance index. PulseDIA-based (DIA [data-independent acquisition]) quantitative proteomics was then employed to reveal the proteome changes in these resistant cells. In total, 7082 proteins from 98,232 peptides were identified and quantified from the dataset using four DIA software tools including OpenSWATH, Spectronaut, DIA-NN, and EncyclopeDIA. Sirtuin signaling pathway was found to be significantly enriched in both ADR-resistant and IMA-resistant K562 cells. In particular, isocitrate dehydrogenase (NADP(+)) 2 was identified as a potential drug target correlated with the drug resistance phenotype, and its inhibition by the antagonist AGI-6780 reversed the acquired resistance in K562 cells to either ADR or IMA. Together, our study has implicated isocitrate dehydrogenase (NADP(+)) 2 as a potential target that can be therapeutically leveraged to alleviate the drug resistance in K562 cells when treated with IMA and ADR., Graphical Abstract, Highlights • Temporal proteomic dynamics in the imatinib or adriamycin-induced drug resistance. • Comparison of four DIA software tools (OpenSWATH, Spectronaut, DIA-NN, and EncyclopeDIA). • Sirtuin signaling pathway was significantly regulated in resistant K562 cells. • IDH2 was identified as a potential drug target correlated for resistant K562 cells., In Brief To understand the underlying resistance mechanisms in response to imatinib (IMA) and adriamycin (ADR), we explored two unique drug resistance models of K562 cells. We applied an optimized DIA–MS method to quantify 98,232 peptides from 7082 proteotypic proteins from these samples using four DIA software tools including OpenSWATH, Spectronaut, DIA-NN, and EncyclopeDIA. The sirtuin signaling pathway was found significantly regulated in both models, and IDH2 was identified as a druggable regulator of acquired drug resistance.

Peptide dendrimers have a broad application in biomedical science due to their biocompatibility, diversity, and multifunctionality, but the precision synthesis of high-molecule weight peptide dendrimers remains challenging. We here report the facile and liquid-phase synthesis of molecular level precision and amino-acid built-in polylysine (PLL) dendrimers with molecular weights as high as ∼60 kDa. Three types of polyhedral oligosilsesquioxane (POSS)-cored PLL dendrimers with phenylalanine, tyrosine, or histidine as building blocks were synthesized. The precise structures of the dendrimers were confirmed by MALDI-TOF MS, GPC, and 1H NMR spectroscopy. The interior functionalized peptide dendrimers improved the encapsulation capability of SN38 and sustained the release profiles. Enhanced molecular interactions between the peptide dendrimers and drugs were explored by both NMR experiments and computer simulations. The peptide dendrimer/SN38 formulations showed potent antitumor activity against multiple cancer cell lines. We believe that this strategy can be applied to the synthesis of tailor-made functional peptide dendrimers for drug-specific delivery and other diverse biomedical applications.

Background: Hypericin is the main active ingredient of St. John’s wort, a Chinese herb commonly used for treating depression. Previous studies shown that hypericin can strongly inhibit human cytochrome P450 (CYP) enzyme activities; however, its potential interactions that inhibit human carboxylesterases 2 (hCE2) are unclear. Purpose: This study aimed to investigate the inhibitory effect of hypericin on hCE2. Methods: The inhibition mechanism of hypericin on hCE2 was studied by using N-(2-butyl-1,3-dioxo-2,3-dihydro- 1H-phenalen-6-yl)-2-chloroacetamide (NCEN). The type of inhibition of hypericin on hCE2 and the corresponding inhibition constant (Ki) value were determined. The inhibition of hypericin on hCE2 in living cells was discussed. The risk of herb-drug interactions (HDI) of hypericin in vivo was predicted by estimating the area under the drug concentration-time curve (AUC) in the presence or absence of hypericin. To understand the inhibition mechanism of hypericin on the activity of hCE2 in-depth, molecular docking was performed. Results: The half-maximal inhibitory concentration (IC50) values of hypericin against the hydrolysis of NCEN and irinotecan (CPT-11) were calculated to be 26.59 μM and 112.8 μM, respectively. Hypericin inhibited the hydrolysis of NCEN and CPT-11. Their Ki values were estimated as 10.53 μM and 81.77 μM, respectively. Moreover, hypericin distinctly suppressed hCE2 activity in living cells. In addition, the AUC of hCE2 metabolic drugs with metabolic sites similar to NCEN was estimated to increase by up to 5 % in the presence of hypericin. More importantly, the exposure of CPT-11 in the intestinal epithelium was predicted to increase by 2 % - 69 % following the oral coadministration of hypericin. Further, molecular simulations indicated that hypericin could strongly interact with ASP98, PHE307, and ARG355 to form four hydrogen bonds within hCE2. Conclusion: These findings regarding the combination of hypericin-containing herbs and drugs metabolized by hCE2 are of considerable clinical significance.

Nonalcoholic fatty liver disease (NAFLD), a metabolic disease, has received wide attention worldwide. However, there is no approved effective drug for NAFLD treatment. In the study, HE and Oil Red O staining were employed to detect liver histopathological changes and the accumulation of lipid droplets. Quantitative real-time PCR, Western blot, bioinformatics, luciferase assay, immunofluorescence staining, reactive oxygen species (ROS), and siRNA were used to further elucidate the mechanism of isoliquiritigenin (ISL) against NAFLD. The results showed that ISL significantly reduced the liver-to-body weight ratios and biochemical index. And the staining results showed that ISL remarkedly ameliorated liver histopathological changes of NAFLD. Furthermore, ISL significantly increased the levels of PPARα, CPT1α, and ACADS, which were involved in lipid metabolism, and inhibited the ROS, TNF-α, IL-1β, and IL-6 expression by activating PGC-1α. Bioinformatics and luciferase assay analysis confirmed that miR-138-5p might bind to PGC-1α mRNA in NAFLD. Importantly, the expression of miR-138-5p was increased in the NAFLD, which was significantly decreased by ISL. In addition, the miR-138-5p inhibitor also promoted lipid metabolism and inhibited inflammatory response in NAFLD via PGC-1α activation. The above results demonstrate that ISL alleviates NAFLD through modulating miR-138-5p/PGC-1α-mediated lipid metabolism and inflammatory reaction in vivo and in vitro.

Polymeric nanocapsules hold considerable applications in cancer drug delivery, but the synthesis of well-defined nanocapsules with a tunable drug release property remains a significant challenge in fabrication. Herein, we demonstrate a supramolecular complexation strategy to assemble small molecular platinum (Pt) compounds into well-defined nanocapsules with high drug loading, acidity-sensitivity, and tunable Pt releasing profile. The design utilizes poly(ethylene glycol)-dendritic polylysine-G4/amides to complex with Pt compounds, forming stable nanocapsules with diameters approximately ∼20 nm and membrane thickness around several nanometers. The stability, drug content, and release profiles are tunable by tailoring the dendritic structure. The designated polymer-Pt nanocapsules, PEG-G4/MSA-Pt, showed sustained blood retention, preferential tumor accumulation, enhanced cellular uptake, lysosomal drug release, and nuclear delivery capability. PEG-G4/MSA-Pt showed enhanced antitumor efficacy compared to free cisplatin and other nanocapsules, which stopped the progression of both A549 cell xenografts and patient-derived xenografts (PDXs) of hepatocellular carcinoma on a mice tumor model. Thus, we believe this strategy is promising for developing Pt-based nanomedicine for cancer drug delivery.

The aim of this study was to explore whether rhein could enhance the effects of pemetrexed (PTX) on the therapy of non-small-cell lung cancer (NSCLC) and to clarify the associated molecular mechanism. Our study shows that rhein in combination with PTX could obviously increase the systemic exposure of PTX in rats, which would be mediated by the inhibition of organic anion transporters (OATs). Furthermore, the toxicity of PTX was significantly raised by rhein in A549 cells in a concentration-dependent manner. Concomitant administration of rhein and PTX-induced cell apoptosis compared with PTX alone in flow cytometry assays, which was further validated by the protein expressions of the apoptotic markers B-cell lymphoma-2/Bcl-2-associated x (Bcl-2/Bax) and Cleaved-Caspase3 (Cl-Caspase3). Meanwhile, the results of monodansylcadaverine (MDC) dyeing experiments showed that PTX-induced autophagy could be enhanced by combination therapy with rhein in A549 cells. Western blot analysis indicated that the synergistic effect of rhein on PTX-mediated autophagy may be interrelated to PI3K-AKT-mTOR pathway inhibition and to the enhancement of p-AMPK and light chain 3-II (LC3-II) protein levels. From these findings, it could be surmised that rhein enhanced the antitumor activity of PTX through influencing autophagy and apoptosis by modulating the PI3K-AKT-mTOR pathway and Bcl-2 family of proteins in A549 cells. Our findings demonstrated that the potential application of rhein as a candidate drug in combination with PTX is promising for treatment of the human lung cancer.

The cyclic dipeptides generally present lower affinity toward intestinal oligopeptide transporter 1 (PEPT1) than the linear dipeptides. JBP485 (cyclo(l-Hyp-l-Ser)) is a low-affinity substrate of PEPT1 with poor oral bioavailability. However, JBP923 (l-Hyp-l-Ser) is a high-affinity substrate of PEPT1 with high oral absorption. We hypothesize that the bioactivatable pseudo-tripeptidization prodrug strategy is promising to increase the affinity of cyclic dipeptides toward PEPT1. To test our hypothesis, we design five amino acid ester prodrugs of JBP485. Compared with JBP485, the optimal prodrug (JBP485-3-CH2-O-valine, J3V) demonstrates improved affinity of PEPT1, oral bioavailability in rats and beagle dogs. Moreover, J3V can dose-dependently protect against liver injury. Additionally, J3V is stable in the gastrointestinal tract, beneficial to the PEPT1-mediated membrane transport, and is bioactivated in the enterocytes and hepatic cells, essential to elicit its bioactivity. In summary, the bioactivatable pseudo-tripeptidization strategy shows potential in increasing affinity of PEPT1 to enhance oral bioavailability of cyclic dipeptides.

Osteosarcoma (OS) is the primary malignant bone tumor that most commonly affects children, adolescents, and young adults. MicroRNA-34a (miR-34a) is involved in tumor metastasis and may be a prognostic marker for patients with cancer. The aim of the present study was to explore the role of miR-34a in patients with OS. The underlying associations between miR-34a expressions and metastasis, recurrence as well as and prognosis were comprehensively analyzed in OS patients.Reverse transcriptase quantitative PCR (RT-qPCR) was used to investigate serum level of miR-34a between clinical OS patients (n = 162) and age-matched healthy controls (n = 162). Expression of miR-34a in OS tissues and adjacent tissues was analyzed using RT-qPCR. RT-qPCR was used to compare the serum level of miR-34a in patients with OS before and after chemotherapy. Multivariate Cox-regression analysis was used to analyze the association between serum level of miR-34a and chemotherapy resistance, overall survival, as well as recurrence and prognosis of OS patients. Five-year recurrence and survival were estimated using Kaplan-Meier curves.Serum level of miR-34a was downregulated in OS patients (n = 86) compared to age-matched healthy controls (n = 86). Expression of miR-34a was downregulated in OS tissue compared to adjacent tissues in clinical patients. The expression of serum miR-34a before and after chemotherapy was positively correlated with the expression of miR-34a in the corresponding tissues. Expression of miR-34a was higher in the group where chemotherapy was effective than that patient where chemotherapy was ineffective. Expression of miR-34a was negatively associated with chemotherapy resistance of OS patients. High serum levels of miR-34a were associated with longer overall survival in OS patients and lower metastasis. Multivariate Cox-regression analysis identified miR-34a serum level with potential prognostic significance.The expression level of serum miR-34a in patients with OS is closely related to the chemotherapy resistance, metastasis, recurrence, and survival of osteosarcoma, which can be used as one of the potential biomarkers and prognosis for the treatment of OS patients. Therefore, miR-34a may be a potential molecular for prediction of the efficacy of chemotherapy and prognosis in OS patients.

To illustrate the microstructural and histomorphological characteristics of osteophytes in the human knee osteoarthritis (OA).Tibial plateau specimens, including osteophytes and subchondral trabecular bone (STB) from weight-bearing and non-weight-bearing regions, were obtained from 81 patients with OA after total knee arthroplasty surgery. All the patients had varus deformity of the knee. Micro-CT was applied to evaluate the microstructure characteristics of the tibial plateau, which was segmented into 6 regions of interest (ROIs). After micro-CT scanning, decalcified and undecalcified bone histology were performed to assess histological features and bone remodeling status in these different ROIs.In both medial and lateral plateaus, osteophytes exhibited a less sclerotic microstructure and higher bone remodeling level compared with STB from weight-bearing and non-weight-bearing regions. Moreover, the medial osteophyte tended to have a more sclerotic microarchitecture and a relatively low level of bone metabolism than its lateral counterpart. Osteophyte, which consists of mesenchymal connective tissue cap, cartilage and mineralized bone tissue, demonstrated evident chondrocyte hypertrophy and endochondral ossification in the cartilage-to-bone interface. Compared with the non-weight-bearing region, the weight-bearing region showed more severely deteriorated cartilage, and more sclerotic subchondral bone with high remodeling status.Osteophytes exhibited a sparse microstructure and high bone remodeling level in knee OA, but demonstrating region-dependent variations. In addition, the microstructure, bone metabolism status and pathological alterations of osteochondral complex were distinct between weight-bearing and non-weight-bearing regions in the tibial plateau. Biomechanical stress might play a pivotal role in osteophyte formation and deterioration of osteochondral complex.

In quantitative real-time PCR (qRT-PCR) detection, the stability of reference genes varies with different organs, tissue locations, sex and developmental stages. This study aimed to screen out and determine the optimal panel of reference genes of the intestine in pre- and post-natal rats of different sex. We used qRT-PCR to detect the mRNA expression of six commonly used reference genes (ACTB, GAPDH, HPRT1, B2M, RPLPO and SDHA) in rat intestines at gestational day 21 (GD21) and postnatal week 12 (PW12). Using GeNorm, BestKeeper and NormFinder software comprehensively analyzed the stability of candidate reference genes and screened out stable reference genes. Further, we used the pathological model of prenatal dexamethasone exposure (PDE) to verify the stability of the selected panel of reference genes. Based on the results of the software analysis, the optimal panel of reference genes in the fetal rat intestine was SDHA + ACTB, and the adult rat small intestine and colon were ACTB + HPRT1 and RPLP0 + GAPDH, respectively. There was no significant sex difference in the above results. Besides, in the PDE model, the results were consistent with those under physiological conditions. Therefore, the stability of intestinal reference genes in fetal rats and adult rats was different, and the intestinal reference genes of adult rats were intestinal segments-specific. The selected panel of reference genes was still stable under pathological conditions. This study determined the optimal panel of reference genes of pre- and post-natal rat intestines and provided reliable reference genes for the qRT-PCR analysis of rat intestines.

Puerarin is an isoflavone isolated from Pueraria lobata (Willd.) Ohwi. In the present study, reversal effect and underlying mechanisms of puerarin on multidrug resistance (MDR) were investigated in K562/ADR cells. K562/ADR cells exhibited adriamycin (ADR) resistance and higher levels of MDR1 expression compared with K562 cells. Puerarin enhanced the chemosensitivity of K562/ADR cells and increased the ADR accumulation in K562/ADR cells. The expression levels of MDR1 were down-regulated by puerarin in K562/ADR cells. Luciferase reporter assay further demonstrated the inhibitory effect of puerarin on TNF-α-induced NF-κB activation. The phosphorylation of IκB-α was significantly suppressed by puerarin. In silico docking analyses suggested that puerarin well matched with the active sites of IκB-α. Moreover, a large number of autophagosomes were found in the cytoplasm of K562/ADR cells after puerarin treatment. The significant increase in LC3-II and beclin-1 was also observed, indicating autophagy induction by puerarin in K562/ADR cells. Puerarin induced cell cycle arrest and apoptosis in K562/ADR cells. Finally, puerarin inhibited phosphorylation of Akt and JNK. In conclusion, puerarin-sensitized K562/ADR cells by downregulating MDR1 expression via inhibition of NF-κB pathway and autophagy induction via Akt inhibition.

With the extensive use of antibiotics in medicine, agriculture and food chemistry, the emergence of multi-drug resistant bacteria become more and more frequent and posed great threats to human health and life. So novel antimicrobial agents were urgently needed to defend the resistant bacteria. Jelleine-I was a small antimicrobial peptide (AMP) with eight amino acids in its sequence. It was believed to be an ideal template for developing antimicrobial agents. In the present study, the possible action mode against both gram-negative bacteria and gram-positive bacteria and in vivo antimicrobial activity was explored. Our results showed that Jelleine-I exhibits its antimicrobial activity mainly by disrupting the integrity of the cell membrane, which would not be affected by the conventional resistant mechanism. It also aims at some intracellular targets such as genomic DNA to inhibit the growth of microbes. In addition, the result of in vivo antimicrobial activity experiment showed that Jelleine-I performed a good therapeutic effect toward the mice with Escherichia coli infected peritonitis. Notably, Jelleine-I has negligible cytotoxicity toward the tested mammalian cells, indicating excellent cell selectivity between prokaryotic cells and eurkayotic cells. In summary, our results showed that Jelleine-I would be a potential candidate to be developed as a novel antimicrobial agent.

Pressure cycling technology (PCT)-assisted tissue lysis and digestion have facilitated reproducible and high-throughput proteomic studies of both fresh-frozen (FF) and formalin-fixed paraffin-embedded (FFPE) tissue of biopsy scale for biomarker discovery. Here, we present an improved PCT method accelerating the conventional procedures by about two-fold without sacrificing peptide yield, digestion efficiency, peptide, and protein identification. The time required for processing 16 tissue samples from tissues to peptides is reduced from about 6 to about 3 h. We analyzed peptides prepared from FFPE hepatocellular carcinoma (HCC) tissue samples by the accelerated PCT method using multiple MS acquisition methods, including short-gradient SWATH-MS, PulseDIA-MS, and 10-plex TMT-based shotgun MS. The data showed that up to 8541 protein groups could be reliably quantified from the thus prepared peptide samples. We applied the accelerated sample preparation method to 25 pairs (tumorous and matched benign) of HCC samples followed by a single-shot, 15 min gradient SWATH-MS analysis. An average of 18 453 peptides from 2822 proteins were quantified in at least 20% samples in this cohort, while 1817 proteins were quantified in at least 50% samples. The data not only identified the previously known dysregulated proteins such as MCM7, MAPRE1, and SSRP1 but also discovered promising novel protein markers, including DRAP1 and PRMT5. In summary, we present an accelerated PCT protocol that effectively doubles the throughput of PCT-assisted sample preparation of biopsy-level FF and FFPE samples without compromising protein digestion efficiency, peptide yield, and protein identification.

Indocyanine green (ICG), a safe and clinically approved near-infrared (NIR) dye, was recently explored as a potential photosensitizer due to its excellent photophysical properties. However, ICG tends to form aggregations in physiological solution, causing fluorescence quenching, fast blood clearance and thereby inefficient tumor accumulation. Herein, we report ICG-based nanodrug delivery systems formed by self-assembly of ICG and chemotherapeutic drugs without any excipients for combined chemo- and photo-therapy. Taking advantage of the amphiphilic aromatic structure, ICG readily bounded with hydrophobic aromatic drugs such as SN38 and formed well-dispersible nanoparticles, which reduced its aggregation-induced quenching and thus greatly improved its photodynamic efficiency. The loaded hydrophobic drugs elicited chemotherapy synergizing the photodynamic therapy, giving rise to much enhanced antitumor activity in vitro and in vivo against human glioblastoma cells and breast cancer cells upon NIR irradiation. The work demonstrates the fabrication of readily translational nanoformulations of hydrophobic drugs using amphiphilic drugs.

The deterioration of medic-patient relation in China raises the question about its impact on patients' adherence to medical instructions. This study focuses on how the quantity and quality of medic-patient communication influences their relationship, and how the perceived relationship at individual level, institution level, and society level influences patients' treatment adherence, through a survey conducted in a city of Southern China (N = 597, Response rate = 66%). The results of path analysis show that consulting time is positively associated with patients' relationship with individual doctors and the whole medical system, as well as their satisfaction with hospital. Unpleasant experience of having disputes with medical workers is negatively associated with the medic-patient relationship at all of the three levels and causes conditional nonadherence. The more positive medic-patient relationship perceived by the patients, the more likely they will display unconditional adherence.

A series of 4-arylamido-2-arylaminoprimidines bearing acrylamide pharmacophore were synthesized as potent EGFRT790M/L858R inhibitors among which 9c (IC50 = 0.5872 nM), 9d (IC50 = 2.213 nM), or 9h (IC50 = 12.57 nM) showed more potent anti-EGFRT790M/L858R activity compared with AZD–9291 (IC50 = 20.80 nM) and possessed high SI displaying 307.6, 56.5, or 12.5 for EGFRT790M/L858R over the wild-type respectively. 9h also showed pretty good activity against H 1975 cells with an IC50 of 1.664 μM and exhibited low toxicity against the normal HBE cells (IC50 > 20 μΜ). 9h had moderate selectivity for H 1975 over A 431 (SI = 7.0) and the other selected cell lines. Morphological staining results further indicated that 9h could promote apoptosis. Hence, 9h was a promising compound for further investigation as a potential EGFRT790M/L858R inhibitor for the treatment of NSCLC.

Germination is an effective biofortification strategy to enhance micronutrients of staple grains. The current study targeted at comparing the effects of normoxia and hypoxia on γ-aminobutyric acid (GABA) content and the componential changes underlying the technofunctionality of germinated wheat. Compared with ungerminated wheat, GABA maximumly enhanced by 88.73% and 50.70% in hypoxia- and normoxia-germinated wheat (HGW and NGW), respectively. Hypoxia suppressed the germination degree of wheat, which better preserved the technofunctionality. Compared with NGW, the proteolytic and α-amylase activities were partially inhibited for HGW, resulting in higher protein and starch content. The diminishment of gluten marcopolymers and soluble high-molecular-weight proteins, and conversion of α-helices to β-turns were alleviated in HGW, which contributed to the more viscoelastic dough. Meanwhile, superior pasting and gelation properties were mainly due to suppressed α-amylase activity and more starch content in HGW. The results suggested that HGW flour can be exploited as a functional staple ingredient.

Background Patients with hypertension usually have to be treated with Angiotensin Converting Enzyme (ACE) inhibitors, and are therefore more exposed to drug-drug Interactions (DDIs) by various mechanisms, especially the mechanisms based on drug metabolizing enzymes and transporters. Objective This review article focuses on the pharmacokinetic interaction mechanisms with ACE inhibitors based on drug metabolizing enzymes and transporters, and the identification of these underlying mechanisms would help physicians and patients to predict, detect and prevent DDIs. Method To identify the pharmacokinetic interaction mechanisms based on drug metabolizing enzymes and transporters of ACE inhibitors. An electronic search of PubMed, Medline, Science Direct, and Springer-Link database was conducted (from 1950 to December, 2017), using drug interactions, cytochrome P450, carboxylesterase, names of transporters, names of ACE inhibitors and pharmacokinetics as keywords. Results and conclusion Inhibition of metabolizing enzymes and transporter system can markedly alter the concentrations of ACE inhibitors. The genetic polymorphisms in the enzymes in some of the specific isoforms seem to explain the inter-individual differences in ACE inhibitors metabolism, and also the inter-individual variation in the pharmacokinetics of ACE inhibitors may be caused by changes in transporter function. Understanding the knowledge of how ACE inhibitors are metabolized and transported is important in predicting and managing DDIs. The data on the roles of drug metabolizing enzymes and transporters in the DDIs of ACE inhibitors should be studied and the selection of ACE inhibitors should be individualized to prevent DDIs in clinic.

Hyperhomocysteinemia (HHCY) has been recognized as an independent risk factor for atherosclerosis and plays a vital role in the development of atherosclerosis. Catalpol, an iridoid glucoside extracted from the root of Rehmannia glutinosa, can produce anti-inflammatory, anti-oxidant, anti-tumor, and dopaminergic neurons protecting effects. This study aimed to determine the protecting effects of catalpol against homocysteine (HCY)-induced injuries in human aortic endothelial cells (HAECs) and uncover the underlying mechanisms: 1. HAECs were cultured with different concentrations of HCY (3 mM) and catalpol (7.5 μΜ, 15 μΜ, 30 μΜ) for 24 h. (1) The level of MDA and GSH as well as LDH release was measured with colorimetric assay. (2) Reactive oxygen species (ROS) were detected by flow cytometry analysis. (3) Western blotting analysis was performed to detect the expression of Nox4, p22phox, ICAM-1, MCP-1, VCAM-1, IκB, nucleus p65, p65 phosphorylation, caspase-3, −9, bax, bcl-2, and ER stress-related proteins. (4) The expressions of CHOP, ATF4 were measured by qRT-PCR. (5) Mitochondrial membrane potential in HCY-treated HAECs was measured by rhodamine 123 staining, and the samples were observed by confocal laser scanning microscopy. 2. DPI, PDTC, and TUDCA were used to determine the interaction among Nox4/ROS, NF-κB, and endoplasmic reticulum stress. 3. TUDCA or Nox4 siRNA were used to investigate whether the effect of catalpol inhibiting the over-production of ROS were associated with inhibiting ER stress and Nox4 expression. Catalpol significantly suppressed LDH release, MDA level, and the reduction of GSH. Catalpol reduced HCY-stimulated ROS over-generation, inhibited the NF-κB transcriptional activation as well as the protein over-expressions of Nox4, ICAM-1, VCAM-1, and MCP-1. Catalpol elevated bcl-2 protein expression and reduced bax, caspase-3, −9 protein expressions in the HCY-treated HAECs. Simultaneously, catalpol could also inhibit the activation of ER stress-associated sensors GRP78, IRE1α, ATF6, P-PERK, P-eIF2α, CHOP, and ATF4 induced by HCY. In addition, the extent of catalpol inhibiting ROS over-generation and NF-κB signaling pathway was reduced after inhibiting Nox4 or ER stress with DPI or TUDCA. The inhibitor of NF-κB PDTC also reduced the effects of catalpol inhibiting the expressions of Nox4 and GRP78. Furthermore, the effect of catalpol inhibiting the over-generation of ROS was reduced by Nox4 siRNA. Catalpol could ameliorate HCY-induced oxidation, cells apoptosis and inflammation in HAECs possibly by inhibiting Nox4/NF-κB and ER stress.

A new class of thiodiphenylpyrimidine analogs (Thio-DPPY) were synthesized as potent and selective EGFR T790M inhibitors to overcome gefitinib resistance in nonsmall cell lung cancer (NSCLC). This structural optimization led to the identification of two potent EGFRT790M/L858R inhibitors, 14a and 14e, which possess IC50 values of 27.5 and 9.1 nM, respectively. Moreover, compounds 14a (SI > 36.4) and 14e (SI > 109.9) exhibited high selectivity and low activity against the wild-type EGFR (IC50 > 1,000 nM). In particular, compound 14a also displayed strong potency against EGFRT790M -mutated H1975 cells (IC50 = 0.074 μM), but weak activity toward normal cells HBE (IC50 > 40 μM) and LO-2 (IC50 = 9.891 μM). It is important that compound 14a (SI = 52.6) significantly improved the selectivity against mutant H1975 cells over wild-type A431 cells than rociletinib (SI = 6.0), thus revealing its slight cell cytotoxicity. This study provides a promising Thio-DPPY derivative as enhanced EGFR T790M inhibitor, and also revealed valuable clues for further optimization of DPPY scaffold to overcome NSCLC resistance.

Acute liver injury (ALI) is a dramatic liver disease characterized by large areas of inflammation in the liver. This study aimed to investigate the protective effects of ginsenoside Rg1 (Rg1), a biologically active component in Panax ginseng, on lipopolysaccharide/ d -galactosamine (LPS/D-GalN)-induced ALI in mice, and meanwhile explore the molecular mechanism in vivo and in vitro. Mice were pretreated with Rg1 for three days prior to LPS (40 μg/kg)/D-GalN (700 mg/kg) administration. The results showed that Rg1 improved the survival rate and reduced the liver to body weight ratios in mice. Rg1 also reduced the production of oxidative markers such as MDA and MPO induced by LPS/D-GalN. In addition, Rg1 significantly decreased the production of inflammatory cytokines including TNF-α, IL-6, IL-1β, Mip-2, Mcp-1, iNOS, and increased the activity of anti-inflammatory cytokine IL-10. Moreover, Rg1 inhibited the protein expression of TLR4 and its downstream genes including NF-κB and MAPKs, which are involved in inflammatory response. Rg1 dramatically reduced oxidative stress by regulating the expression of efflux transporters Mrp2 and various enzymes including GCLC, GCLM, HO-1 and NQO1. However, the changes in these genes and protein induced by Rg1 were abrogated by TLR4 antagonist TAK-242 in vitro. In conclusion, Rg1 had hepatoprotective effect on LPS/D-GalN-induced ALI in mice. The protection may be associated with the inhibition of TLR4. These findings suggest that Rg1 may be a promising agent for prevention against ALI.

Cholestasis is a clinical syndrome with systemic and intrahepatic accumulation of excessive toxic bile acids that ultimately cause hepatobiliary injury. Recently obeticholic acid (OCA) which is a farnesoid X receptor (FXR) agonist was approved by FDA to treat cholestatic liver diseases, which provided us a newly therapeutic strategy against cholestasis. The purpose of the current study is to screen novel FXR agonists and verify the anti-cholestasis effect of yangonin in vivo and in vitro. The computational strategy of two-dimensional virtual screening was used to search for new FXR agonists, and dual-luciferase reporter gene assay was used to further demonstrate FXR activation by yangonin. Then, the hepatoprotective effect of yangonin via FXR activation against cholestasis and hepatotoxity was evaluated in mice and was investigated using FXR silence in cells. Yangonin was found to activate FXR to exert hepatoprotective effect against cholestatic liver injury. Dynamic change analysis of bile acids and gene analysis revealed that yangonin promoted bile acid efflux into bile and reduced hepatic uptake via the regulation of FXR-target genes Bsep, Mrp2 and Ntcp expression. Furthermore, yangonin modulated enzymes involved in bile acid synthesis and metabolism including Cyp7a1 Cyp8b1 and Sult2a1. In addition, yangonin promoted liver repair and suppressed liver inflammation. However, the changes in these genes and protein, as well as ameliorative liver histology induced by yangonin were abrogated by FXR antagonist guggulsterone in vivo and FXR siRNA in vitro. Yangonin produces protective effect against cholestasis via FXR activation. Yangonin may be an effective approach for the prevention and treatment for cholestatic liver diseases.

Hepatic fibrosis is a pathological process that eventually leads to the development of cirrhosis and liver cancer by various types of chronic liver disease. To date, there is no standard treatment for the progression of liver fibrosis. This study aims to investigate the hepatoprotection of auraptene (AUR), a simple coumarin contained in the peels of citrus fruits such as grapefruit, against thioacetamide (TAA)-induced hepatic fibrosis in mice. The involvement of farnesoid X receptor (FXR) in the anti-fibrotic effect of AUR was further elucidated using in vivo and in vitro experiments. AUR was found to remarkably protect against liver injury induced by TAA in mice and maintain the homeostasis of bile acids via the regulation of FXR-target genes including Bsep, Mrp2, Ntcp, Cyp7a1 and Cyp8b1. Masson and Sirius red staining indicated a reduction of the collagen content in the liver of AUR treated mice. Furthermore, AUR inhibited the activation of hepatic stellate cells (HSCs) by down-regulating the expression of TGF-β1 and α-SMA and expressed anti-inflammatory effects by reducing the expression of NF-κB, TNF-α and IL-1β. However, the changes in these genes and protein as well as ameliorative liver histology induced by AUR were abrogated by FXR antagonist guggulsterone in vivo and FXR siRNA in vitro. Overall, AUR protects against TAA-induced hepatic fibrosis due to the reduction of toxic bile acids and inhibition of hepatic stellate cell (HSC) activation and inflammation, which were all in association with FXR activation. AUR might be efficacious for the prevention and treatment of hepatic fibrosis in mice.

The BTK and JAK3 receptor tyrosine kinases are two validated and therapeutically amenable targets in the treatment of B-cell lymphomas. Here we report the identification of several classes of pyrimidine derivatives as potent BTK and JAK3 dual inhibitors. Among these molecules, approximately two thirds displayed strong inhibitory capacity at less than 10 nM concentration, and four compounds (7e, 7g, 7m and 7n) could significantly inhibit the phosphorylation of BTK and JAK3 enzymes at concentrations lower than 1 nM. Additionally, these pyrimidine derivatives also exhibited enhanced activity to block the proliferation of B-cell lymphoma cells compared with the representative BTK inhibitor ibrutinib. In particular, two structure-specific compounds 7b and 7e displayed stronger activity than reference agents in cell-based evaluation, with IC50 values lower than 10 μM. Further biological studies, including flow cytometric analysis, and a xenograft model for in vivo evaluation, also indicated their efficacy and low toxicity in the treatment of B-cell lymphoma. These findings provide a new insight for the development of novel anti-B-cell lymphoma drugs with multi-target actions.

Due to the threat of escalating multi-drug resistant gram-negative bacteria to human health and life, novel antimicrobial agents against gram-negative pathogens are urgently needed. As antimicrobial peptides are not prone to induce bacteria resistance, they are believed to be one kind of promising class of potential antimicrobial agent candidates to combat multi-drug resistant bacteria for long-term use. Jelleine-1, first isolated from the royal jelly of honeybees, is a typical amphiphilic antimicrobial peptide and shows broad antimicrobial spectrum and negligible toxicity. To promote its antimicrobial activity and extend its potential of clinical use against multi-drug resistant gram-negative bacteria, novel analogs of jelleine-1 were designed, synthesized and their antimicrobial functions and toxicity were examined in this study. Our results showed that fine tuning of the cationic charge, polarity, and basicity of the sequence through amino acids substitution at position 3, 5, 7 and maintaining position 1, 4, 6, 8 unchanged could improve the bioactivity of jelleine-1 significantly. Meanwhile, we also found that the substitution of phenylalanine by tryptophan also could improve the antimicrobial activity of jelleine-1. Among all the analogs, analog 15, which is enriched in arginine and leucine, showed the most potent antimicrobial activity against both gram-negative and gram-positive bacteria, especially to multi-drug resistant Pseudomonas aeruginosa in vivo and in vitro. In addition, analog 15 also showed potent inhibition of the formation of multi-drug resistant P. aeruginosa biofilm and negligible toxicity, which was certified by MTT, hemolysis, blood assay, and biochemical analysis.

Cilostazol undergoes extensive liver metabolism. However, the transporter-mediated hepatic disposition of cilostazol remains unknown. The present study was performed to investigate the hepatic uptake and biliary excretion of cilostazol and its metabolites (OPC-13015 and OPC-13213) using rat liver and human transporter-transfected cells in vitro. Cilostazol uptake by rat liver slices and isolated rat hepatocytes exhibited time-, concentration-, and temperature dependency and was decreased by Oatp inhibitors, which suggested that Oatp was involved in the hepatic uptake of cilostazol. Cilostazol uptake in rat hepatocytes, OATP1B1-, and OATP1B3-HEK293 cells indicated a saturable process with Km values of 2.7 μM, 17.7 μM, and 2.7 μM, respectively. Epigallocatechin gallate, cyclosporin A, rifampicin, and telmisartan inhibited cilostazol uptake in OATP1B1/1B3-HEK293 cells with Ki values close to their clinical plasma concentration, which suggested possible drug-drug interactions in humans via OATP1B1/1B3. Moreover, the cumulative biliary excretion of cilostazol and OPC-13015 was significantly decreased by quinidine, bilirubin, and novobiocin in perfused rat liver, but OPC-13213 biliary excretion was only inhibited by novobiocin, which suggested that the efflux transporters Mrp2, Bcrp, and P-gp were involved in the biliary excretion of cilostazol and its metabolites. Our findings indicated that multiple transporters were involved in the hepatic disposition of cilostazol and its metabolites.

A class of sulfonamide-substituted diphenylpyrimidines (Sul-DPPYs) were synthesized to improve activity against the focal adhesion kinase (FAK). Most of these new Sul-DPPYs displayed moderate activity against the FAK enzyme with IC50 values of less than 100nM; regardless, they could effectively inhibit several classes of refractory cancer cell lines with IC50 values of less than 10µM, including the pancreatic cancer cell lines (AsPC-1, Panc-1 and BxPC-3), the NSCLC-resistant H1975 cell line, and the B lymphocyte cell line (Ramos cells). Results of flow cytometry indicated that inhibitor 7e promoted apoptosis of pancreatic cancer cells in a dose-dependent manner. In addition, it almost completely induced the apoptosis at a concentration of 10µM. Compound 7e may be selected as a potent FAK inhibitor for the treatment of pancreatic cancer.

We investigated the reversal effect of afatinib (AFT) on activity of adriamycin (ADR) in A549T cells and clarified the related molecular mechanisms. A549T cells overexpressing P-glycoprotein (P-gp) were resistant to anticancer drug ADR. AFT significantly increased the antitumor activity of ADR in A549T cells. AFT increased the intracellular concentration of ADR by inhibiting the function and expression of P-gp at mRNA and protein levels in A549T cells. Additionally, the reversal effect of AFT on P-gp mediated multidrug resistance (MDR) might be related to the inhibition of PI3K/Akt pathway. Cotreatment with AFT and ADR could enhance ADR-induced apoptosis and autophagy in A549T cells. Meanwhile, the co-treatment significantly induced cell apoptosis and autophagy accompanied by increased expression of cleaved caspase-3, PARP, LC3B-II, and beclin 1. Apoptosis inhibitors had no significant effect on cell activity, while autophagy inhibitors decreased cell viability, suggesting that autophagy may be a self protective mechanism of cell survival in the absence of chemotherapy drugs. Interestingly, when combined with AFT and ADR, inhibition of apoptosis and/or autophagy could enhance cell viability. These results indicated that in addition to inhibit P-gp, ADR-induced apoptosis, and autophagy promoted by AFT contributed to the antiproliferation effect of combined AFT and ADR on A549T cells. These findings provide evidence that AFT combined ADR may achieve a better therapeutic effect to lung cancer in clinic. J. Cell. Biochem. 119: 414-423, 2018. © 2017 Wiley Periodicals, Inc.

Potential new EGFRT790M inhibitors comprised of structurally modified diphenylpyrimidine derivatives bearing a morpholine functionality (Mor-DPPYs) were used to improve the activity and selectivity of gefitinib-resistant non-small cell lung cancer (NSCLC) treatment. This led to the identification of inhibitor 10c, which displayed high activity against EGFRT790M/L858R kinase (IC50 = 0.71 nM) and repressed H1975 cell replication harboring EGFRT790M mutations at a concentration of 0.037 μM. Inhibitor 10c demonstrated high selectivity (SI = 631.9) for T790M-containing EGFR mutants over wild type EGFR, suggesting that it will cause less side effects. Moreover, this compound also shows promising antitumor efficacy in a murine EGFRT790M/L858R-driven H1975 xenograft model without affecting body weight. This study provides new potential lead compounds for further development of anti-NSCLC drugs.

A new series of diphenylpyrimidine derivatives (DPPYs) bearing various aniline side chains at the C-2 position of pyrimidine core were synthesized as potent BTK inhibitors. Most of these inhibitors displayed improved activity against B leukemia cell lines compared with lead compound spebrutinib. Subsequent studies showed that the peculiar inhibitor 7j, with IC50 values of 10.5 μM against Ramos cells and 19.1 μM against Raji cells, also displayed slightly higher inhibitory ability than the novel agent ibrutinib. Moreover, compound 7j is not sensitive to normal cells PBMC, indicating low cell cytotoxicity. In addition, flow cytometry analysis indicated that 7j significantly induced the apoptosis of Ramos cells, and arrested the cell cycle at the G0/G1 phase. These explorations provided new clues to discover pyrimidine scaffold as more effective BTK inhibitors.

Idiopathic pulmonary fibrosis (IPF) is a serious and fatal lung disease, with a median survival of only 3–5 years from diagnosis. Janus kinase 3 (JAK3) has a well-established role in the pathogenesis of various autoimmune diseases, including rheumatoid arthritis (RA) and autoimmune-related pulmonary fibrosis. In this study, through the use of a conformationally-constrained design strategy, a series of thieno[3,2-d]pyrimidines were synthesized as potent JAK3 inhibitors for the treatment of IPF. Among them, the most potent JAK3 inhibitor, namely 8e (IC50 = 1.38 nM), significantly reduced the degree of airsacculitis and fibrosis according to hematoxylin-eosin (HE) staining assay for the lung tissue in the bleomycin (BLM)-induced pulmonary fibrosis mouse model. The clear reduction of the lung collagen deposition by the determination of Masson and hydroxyproline (HYP) content also demonstrated its efficacy in the treatment of fibrosis. In addition, 8e also reduced the expression of the inflammatory markers IL-6, IL-17A, TNF-α and malondialdehyde (MDA) in lung tissue, which indicated its higher anti-inflammatory activity compared with that of the reference agents (nintedanib and gefitinib). Furthermore, it possessed low cytotoxicity against normal human bronchial epithelia (HBE) cells (IC50 > 39.0 μM) and C57BL mice. All these evaluated biological properties suggest that 8e may be a potential JAK3 inhibitor for the treatment of IPF.

JAK3 is predominantly expressed in hematopoietic cells and has been a promising therapeutic target for the treatment of B-cell lymphoma. In this study, a new class of thieno[3,2-d]pyrimidines harboring acrylamide pharmacophore were synthesized as potent covalent JAK3 inhibitors (IC50

This study was devised to investigate if P-glycoprotein (P-gp) mediated the drug–drug interaction (DDI) between genistein and repaglinide. When genistein was added, the plasma concentrations of repaglinide in rats were increased. The maximum plasma concentration (Cmax) of repaglinide increased from 70.80 ± 7.98 ng/mL to 124.71 ± 9.02 ng/mL and the area under the plasma concentration-time curve (AUC) increased from 134.89 ± 13.65 μg·h/L to 245.95 ± 7.24 μg·h/L. Intestinal absorption of repaglinide was markedly enhanced by genistein or P-gp inhibitor verapamil (Ver), both in situ rat jejunal perfusion studies and in vitro transport assays using everted rat intestinal sac preparations. Furthermore, the accumulation of repaglinide in both Caco-2 cells and IEC-6 cells also increased significantly in the presence of genistein and Ver. The transepithelial transport rate of repaglinide from basolateral-to-apical in MDR1-MDCK cells was 3.6-fold higher than the apical-to-basolateral rate with a net efflux ratio of 1.92 compared with mock-MDCK cells, which was significantly decreased following co-administration with genistein or Ver. In an intracellular accumulation experiment using Rhodamine 123 as a P-gp substrate, genistein significantly increased the intracellular fluorescence of Rhodamine 123. These results indicated that genistein had an inhibitory effect on the efflux function of P-gp. Through molecular docking assays we further found that genistein could bind to the nucleotide-binding domains (NBD) in the cytoplasm of P-gp, thus affecting the functions of P-gp. In conclusion, genistein inhibited the efflux of repaglinide mediated by P-gp in rats and in vitro. The findings suggested that the DDI between genistein and repaglinide is mediated by P-gp, and a dosage adjustment may be needed when they are co-administered in a clinical setting.

A new class of pyrimidine derivatives were designed and synthesized as potential dual FAK and EGFRT790M inhibitors using a fragment-based drug design strategy. This effort led to the identification of the two most active inhibitors, namely 9a and 9f, against both FAK (IC50 = 1.03 and 3.05 nM, respectively) and EGFRT790M (IC50 = 3.89 and 7.13 nM, respectively) kinase activity. Moreover, most of these compounds also exhibited strong antiproliferative activity against the three evaluated FAK-overexpressing pancreatic cancer (PC) cells (AsPC-1, BxPC-3, Panc-1) and two drug-resistant cancer cell lines (breast cancer MCF-7/adr cells and lung cancer H1975 cells) at concentrations lower than 6.936 μM. In addition, 9a was also effective in the in vivo assessment conducted in a FAK-driven human AsPC-1 cell xenograft mouse model. Overall, this study offers a new insight into the treatment of hard to treat cancers.

Background Our previous study revealed that microRNA-125a-5p plays a crucial role in regulating hepatic glycolipid metabolism by targeting STAT3 in type 2 diabetes mellitus (T2DM). Dioscin, a major active ingredient in Dioscoreae nipponicae rhizomes, displays various pharmacological activities, but its role in T2DM has not been reported. Purpose The aim of this study was to investigate the effect of dioscin on T2DM and elucidate its potential mechanism. Methods The effect of dioscin on glycolipid metabolic disorder in insulin-induced HepG2 cells, palmitic acid-induced AML12 cells, high-fat diet- and streptozotocin- induced T2DM rats, and spontaneous T2DM KK-Ay mice were evaluated. Then, the possible mechanisms of dioscin were comprehensively evaluated. Results Dioscin markedly alleviated the dysregulation of glycolipid metabolism in T2DM by reducing hyperglycemia and hyperlipidemia, improving insulin resistance, increasing hepatic glycogen content, and attenuating lipid accumulation. When the mechanism was investigated, dioscin was found to markedly elevate miR-125a-5p level and decrease STAT3 expression. Consequently, dioscin increased phosphorylation levels of STAT3, PI3K, AKT, GSK-3β, and FoxO1 and decreased gene levels of PEPCK, G6Pase, SREBP-1c, FAS, ACC, and SCD1, leading to an increase in glycogen synthesis and a decrease in gluconeogenesis and lipogenesis. The effects of dioscin on regulating miR-125a-5p/STAT3 pathway were verified by miR-125a-5p overexpression and STAT3 overexpression. Conclusions Dioscin showed potent anti-T2DM activity by improving the inhibitory effect of miR-125a-5p on STAT3 signaling to alleviate glycolipid metabolic disorder of T2DM.