Mechanistic Understanding of Dexamethasone-Mediated Protection against Remdesivir-Induced Hepatotoxicity.

Remdesivir (RDV), a broad-spectrum antiviral agent, is often used together with dexamethasone (DEX) for hospitalized COVID-19 patients requiring respiratory support. Potential hepatic adverse drug reaction is a safety concern associated with the use of RDV. We previously reported that DEX cotreatment effectively mitigates RDV-induced hepatotoxicity and reduces elevated serum alanine aminotransferase and aspartate aminotransferase levels in cultured human primary hepatocytes (HPH) and hospitalized COVID-19 patients, respectively. Yet, the precise mechanism behind this protective drug-drug interaction remains largely unknown. Here, we show that through the activation of p38, c-Jun N-terminal kinase (JNK), and extracellular signal-regulated kinases 1 and 2 (ERK1/2) signaling, RDV induces apoptosis (cleavage of caspases 8, 9, and 3), autophagy (increased autophagosome and LC3-II), and mitochondrial damages (decreased membrane potential, respiration, ATP levels, and increased expression of Bax and the released cytosolic cytochrome C) in HPH. Importantly, cotreatment with DEX partially reversed RDV-induced apoptosis, autophagy, and cell death. Mechanistically, DEX deactivates/dephosphorylates p38, JNK, and ERK1/2 signaling by enhancing the expression of dual specificity protein phosphatase 1 (DUSP1), a mitogen-activated protein kinase (MAPK) phosphatase, in a glucocorticoid receptor (GR)-dependent manner. Knockdown of GR in HPH attenuates DEX-mediated DUSP1 induction, MAPK dephosphorylation, as well as protection against RDV-induced hepatotoxicity. Collectively, our findings suggest a molecular mechanism by which DEX modulates the GR-DUSP1-MAPK regulatory axis to alleviate the adverse actions of RDV in the liver. SIGNIFICANCE STATEMENT: The research uncovers the molecular mechanisms by which dexamethasone safeguards against remdesivir-associated liver damage in the context of COVID-19 treatment.

Synergistic effects of graphene microgrooves and electrical stimulation on M2 macrophage polarization.

Macrophages play a crucial role in host response and wound healing, with M2 polarization contributing to the reduction of foreign-body reactions induced by the implantation of biomaterials and promoting tissue regeneration. Electrical stimulation (ES) and micropatterned substrates have a significant impact on the macrophage polarization. However, there is currently a lack of well-established cell culture platforms for studying the synergistic effects of these two factors. In this study, we prepared a graphene free-standing substrate with 20 µm microgrooves using capillary forces induced by water evaporation. Subsequently, we established an ES cell culture platform for macrophage cultivation by integrating a self-designed multi-well chamber cell culture device. We observed that graphene microgrooves, in combination with ES, significantly reduce cell spreading area and circularity. Results from immunofluorescence, ELISA, and flow cytometry demonstrate that the synergistic effect of graphene microgrooves and ES effectively promotes macrophage M2 phenotypic polarization. Finally, RNA sequencing results reveal that the synergistic effects of ES and graphene microgrooves inhibit the macrophage actin polymerization and the downstream PI3K signaling pathway, thereby influencing the phenotypic transition. Our results demonstrate the potential of graphene-based microgrooves and ES to synergistically modulate macrophage polarization, offering promising applications in regenerative medicine.

Degradable Antimicrobial Ureteral Stent Construction with Silver@graphdiyne Nanocomposite.

In the surgical treatment of urinary diseases, ureteral stents are commonly used interventional medical devices. Although polymer ureteral stents with polyurethane as the main constituent are widely used in the clinic, the need for secondary surgery to remove them and their propensity to cause bacterial infections greatly limit their effectiveness. To satisfy clinical requirements, an electrospinning-based strategy to fabricate PLGA ureteral stents with silver@graphdiyne is innovated. Silver (Ag) nanoparticles are uniformly loaded on the surface of graphdiyne (GDY) flakes. It is found that the incorporation of Ag nanoparticles into GDY markedly increases their antibacterial properties. Subsequently, the synthesized and purified Ag@GDY is homogeneously blended with poly(lactic-co-glycolic acid) (PLGA) as an antimicrobial agent, and electrospinning along with high-speed collectors is used to make tubular stents. The antibacterial effect of Ag@GDY and the porous microstructure of the stents can effectively prevent bacterial biofilm formation. Furthermore, the stents gradually decrease in toughness but increase in strength during the degradation process. The cellular and subcutaneous implantation experiments demonstrate the moderate biocompatibility of the stents. In summary, considering these performance characteristics and the technical feasibility of the approach taken, this study opens new possibilities for the design and application of biodegradable ureteral stents.

Multi-omics analysis of an in vitro photoaging model and protective effect of umbilical cord mesenchymal stem cell-conditioned medium.

BACKGROUND: Skin ageing caused by long-term ultraviolet (UV) irradiation is a complex biological process that involves multiple signalling pathways. Stem cell-conditioned media is believed to have anti-ageing effects on the skin. The purpose of this study was to explore the biological effects of UVB irradiation and anti-photoaging effects of human umbilical cord mesenchymal stem cell-conditioned medium (hUC-MSC-CM) on HaCaT cells using multi-omics analysis with a novel cellular photoaging model. METHODS: A cellular model of photoaging was constructed by irradiating serum-starved HaCaT cells with 20 mJ/cm2 UVB. Transcriptomics and proteomics analyses were used to explore the biological effects of UVB irradiation on photoaged HaCaT cells. Changes in cell proliferation, apoptosis, and migration, the cell cycle, and expression of senescence genes and proteins were measured to assess the protective effects of hUC-MSC-CM in the cellular photoaging model. RESULTS: The results of the multi-omics analysis revealed that UVB irradiation affected various biological functions of cells, including cell proliferation and the cell cycle, and induced a senescence-associated secretory phenotype. hUC-MSC-CM treatment reduced cell apoptosis, inhibited G1 phase arrest in the cell cycle, reduced the production of reactive oxygen species, and promoted cell motility. The qRT-PCR results indicated that MYC, IL-8, FGF-1, and EREG were key genes involved in the anti-photoaging effects of hUC-MSC-CM. The western blotting results demonstrated that C-FOS, C-JUN, TGFß, p53, FGF-1, and cyclin A2 were key proteins involved in the anti-photoaging effects of hUC-MSC-CM. CONCLUSION: Serum-starved HaCaT cells irradiated with 20 mJ/cm2 UVB were used to generate an innovative cellular photoaging model, and hUC-MSC-CM demonstrates potential as an anti-photoaging treatment for skin.

circPTP4A2-miR-330-5p-PDK2 Signaling Facilitates In Vivo Survival of HuMSCs on SF-SIS Scaffolds and Improves the Repair of Damaged Endometrium.

Background: Human umbilical cord mesenchymal stem cells- (HuMSCs-) based therapy has shown promising results in the treatment of intrauterine adhesions (IUA). In this study, we aimed to construct a HuMSCs-seeded silk fibroin small-intestinal submucosa (SF-SIS) scaffold and evaluate its ability to repair the damaged endometrium in an IUA mouse model. Methods: To identify the functional effect of HuMSCs-SF-SIS scaffolds on the repair of damaged endometrium, a mouse IUA model was established. Uterine morphology and fibrosis were evaluated by hematoxylin-eosin staining and Masson staining. CircRNA sequencing, real-time PCR, and RNA fluorescence in situ hybridization were used to screen and verify the potential circRNAs involved in the repair of damaged endometrium by HuMSCs. Real-time integrated cellular measurement of oxygen consumption rate was performed using the Seahorse XF24 Extracellular Flux Analyzer. The potential downstream miRNAs and proteins of circRNAs were analyzed by dual-luciferase reporter assay and western blot. Results: HuMSCs-SF-SIS not only increased the number of glands but also reduced the ulcer area in the IUA model. circPTP4A2 was elevated in the HuMSCs seeded on the SF-SIS scaffolds and was targeted by miR-330-5p-PDK2. It also stabilized the mitochondrial metabolism of HuMSCs. Moreover, miR-330-5p was found to inhibit PDK2 expression through the 3' UTR target region. A rescue experiment further showed that circPTP4A2-miR-330-5p-PDK2 signaling was critical to HuMSCs-SF-SIS in decreasing the fibrosis area and increasing the number of glands in the IUA model. Conclusion: We demonstrated that circPTP4A2 was elevated in HuMSCs-seeded on SF-SIS scaffolds and stabilized the mitochondrial metabolism through miR-330-5p-PDK2 signaling, which contributes to endometrial repair progression. These findings demonstrate that HuMSCs-seeded SF-SIS scaffolds have potential for the treatment of IUA.

Targeting CAR and Nrf2 improves cyclophosphamide bioactivation while reducing doxorubicin-induced cardiotoxicity in triple-negative breast cancer treatment.

Cyclophosphamide (CPA) and doxorubicin (DOX) are key components of chemotherapy for triple-negative breast cancer (TNBC), although suboptimal outcomes are commonly associated with drug resistance and/or intolerable side effects. Through an approach combining high-throughput screening and chemical modification, we developed CN06 as a dual activator of the constitutive androstane receptor (CAR) and nuclear factor erythroid 2-related factor 2 (Nrf2). CN06 enhances CAR-induced bioactivation of CPA (a prodrug) by provoking hepatic expression of CYP2B6, while repressing DOX-induced cytotoxicity in cardiomyocytes in vitro via stimulating Nrf2-antioxidant signaling. Utilizing a multicellular coculture model incorporating human primary hepatocytes, TNBC cells, and cardiomyocytes, we show that CN06 increased CPA/DOX-mediated TNBC cell death via CAR-dependent CYP2B6 induction and subsequent conversion of CPA to its active metabolite 4-hydroxy-CPA, while protecting against DOX-induced cardiotoxicity by selectively activating Nrf2-antioxidant signaling in cardiomyocytes but not in TNBC cells. Furthermore, CN06 preserves the viability and function of human iPSC-derived cardiomyocytes by modulating antioxidant defenses, decreasing apoptosis, and enhancing the kinetics of contraction and relaxation. Collectively, our findings identify CAR and Nrf2 as potentially novel combined therapeutic targets whereby CN06 holds the potential to improve the efficacy/toxicity ratio of CPA/DOX-containing chemotherapy.

Human constitutive androstane receptor represses liver cancer development and hepatoma cell proliferation by inhibiting erythropoietin signaling.

The constitutive androstane receptor (CAR) is a nuclear receptor that plays a crucial role in regulating xenobiotic metabolism and detoxification, energy homeostasis, and cell proliferation by modulating the transcription of numerous target genes. CAR activation has been established as the mode of action by which phenobarbital-like nongenotoxic carcinogens promote liver tumor formation in rodents. This paradigm, however, appears to be unrelated to the function of human CAR (hCAR) in hepatocellular carcinoma (HCC), which remains poorly understood. Here, we show that hCAR expression is significantly lower in HCC than that in adjacent nontumor tissues and, importantly, reduced hCAR expression is associated with a worse HCC prognosis. We also show overexpression of hCAR in human hepatoma cells (HepG2 and Hep3B) profoundly suppressed cell proliferation, cell cycle progression, soft-agar colony formation, and the growth of xenografts in nude mice. RNA-Seq analysis revealed that the expression of erythropoietin (EPO), a pleiotropic growth factor, was markedly repressed by hCAR in hepatoma cells. Addition of recombinant EPO in HepG2 cells partially rescued hCAR-suppressed cell viability. Mechanistically, we showed that overexpressing hCAR repressed mitogenic EPO-EPO receptor signaling through dephosphorylation of signal transducer and activator of transcription 3, AKT, and extracellular signal-regulated kinase 1/2. Furthermore, we found that hCAR downregulates EPO expression by repressing the expression and activity of hepatocyte nuclear factor 4 alpha, a key transcription factor regulating EPO expression. Collectively, our results suggest that hCAR plays a tumor suppressive role in HCC development, which differs from that of rodent CAR and offers insight into the hCAR-hepatocyte nuclear factor 4 alpha-EPO axis in human liver tumorigenesis.

Modification of polyether ether ketone for the repairing of bone defects.

Bone damage as a consequence of disease or trauma is a common global occurrence. For bone damage treatment-bone implant materials are necessary across three classifications of surgical intervention (i.e. fixation, repair, and replacement). Many types of bone implant materials have been developed to meet the requirements of bone repair. Among them, polyether ether ketone (PEEK) has been considered as one of the next generation of bone implant materials, owing to its advantages related to good biocompatibility, chemical stability, x-ray permeability, elastic modulus comparable to natural bone, as well as the ease of processing and modification. However, as PEEK is a naturally bioinert material, some modification is needed to improve its integration with adjacent bones after implantation. Therefore, it has become a very hot topic of biomaterials research and various strategies for the modification of PEEK including blending, 3D printing, coating, chemical modification and the introduction of bioactive and/or antibacterial substances have been proposed. In this systematic review, the recent advances in modification of PEEK and its application prospect as bone implants are summarized, and the remaining challenges are also discussed.

Surface functionalized biomimetic bioreactors enable the targeted starvation-chemotherapy to glioma.

Altering the glucose supply and the metabolic pathways would be an intriguing strategy in starvation therapy toward cancers. Nevertheless, starvation therapy alone could be inadequate to eliminate tumor cells completely. Herein, a multifunctional bioreactor was fabricated for synergistic starvation-chemotherapy through embedding glucose oxidase (GOx) and doxorubicin (DOX) in the tumor targeting ligands (RGD) modified red blood cell membrane camouflaged metal-organic framework (MOF) nanoparticle (denoted as RGD-mGZD). Owing to the remarkable biointerfacing property, the designed RGD-mGZD could not only possess enhanced blood retention time inherited from red blood cells, but also preferentially target the tumor site after the modification with RGD peptide. Once the bioreactor reached the desired region, GOx promptly consumed the intratumoral glucose and oxygen to starve cancer cells for robust starvation therapy. More importantly, the aggravated acidic microenvironment at the tumor region was found to induce the decomposition of the MOF structure, thus triggering the release of DOX for reinforced chemotherapy. This bioreactor would further prompt the development of synergistic patterns toward cancer treatment in a spatiotemporally controlled manner.

Androgen-induced insulin resistance is ameliorated by deletion of hepatic androgen receptor in females.

Androgen excess is one of the most common endocrine disorders of reproductive-aged women, affecting up to 20% of this population. Women with elevated androgens often exhibit hyperinsulinemia and insulin resistance. The mechanisms of how elevated androgens affect metabolic function are not clear. Hyperandrogenemia in a dihydrotestosterone (DHT)-treated female mouse model induces whole body insulin resistance possibly through activation of the hepatic androgen receptor (AR). We investigated the role of hepatocyte AR in hyperandrogenemia-induced metabolic dysfunction by using several approaches to delete hepatic AR via animal-, cell-, and clinical-based methodologies. We conditionally disrupted hepatocyte AR in female mice developmentally (LivARKO) or acutely by tail vein injection of an adeno-associated virus with a liver-specific promoter for Cre expression in ARfl/fl mice (adLivARKO). We observed normal metabolic function in littermate female Control (ARfl/fl ) and LivARKO (ARfl/fl ; Cre+/- ) mice. Following chronic DHT treatment, female Control mice treated with DHT (Con-DHT) developed impaired glucose tolerance, pyruvate tolerance, and insulin tolerance, not observed in LivARKO mice treated with DHT (LivARKO-DHT). Furthermore, during an euglycemic hyperinsulinemic clamp, the glucose infusion rate was improved in LivARKO-DHT mice compared to Con-DHT mice. Liver from LivARKO, and primary hepatocytes derived from LivARKO, and adLivARKO mice were protected from DHT-induced insulin resistance and increased gluconeogenesis. These data support a paradigm in which elevated androgens in females disrupt metabolic function via hepatic AR and insulin sensitivity was restored by deletion of hepatic AR.

Effects of intermittent fasting on liver physiology and metabolism in mice.

A broad spectrum of health benefits from intermittent fasting have been reported in studies on animal models and human subjects. However, the underlying mechanisms of these beneficial effects remain largely elusive. The present study aimed to explore the effects and potential mode of action of intermittent fasting in mouse models with a focus on the liver. C57BL/6 mice were subjected to intermittent fasting or ad libitum feeding as controls. It was determined that 12 h of daily intermittent fasting for 30 days significantly reduced the cumulative food intake compared with that in mice with ad libitum feeding. Fasting resulted in a significantly reduced liver mass but only had a minimal effect on bodyweight. The effects on the liver by 30 days of fasting were not reversed by subsequent ad libitum refeeding for 30 days. Among the measured blood biochemical parameters, the levels of blood glucose were decreased, while the levels of alkaline phosphatase were increased in fasting mice. Of note, targeted metabolic profiling revealed global elevation of metabolites in the livers of fasting mice. These metabolic molecules included adenosine triphosphate, nicotinamide adenine dinucleotide phosphate (NADP), reduced NADP and succinate, which are essentially involved in the citric acid cycle and oxidative phosphorylation. Thus, it was concluded that daily 12 h of intermittent fasting for one month significantly reduced the liver weight of mice, which is associated with enhanced liver metabolism.

Inside-Out-Oriented Cell Membrane Biomimetic Magnetic Nanoparticles for High-Performance Drug Lead Discovery.

Biomimetic cell membrane-coated nanoparticles have been broadly applied because of their superior biochemical properties. The right-side-out cell membrane coating manner provides nanoparticles with an immune-evasive stealth function in vivo. However, this acts as a drag for drug discovery when the drug targets are the intracellular domain of transmembrane receptors. Herein, inside-out-oriented cell membrane-coated nanoparticles were prepared for screening tyrosine kinase inhibitors, which specifically interacted with the intracellular kinase domain of the epidermal growth factor receptor. Biotinylated human lung adenocarcinoma epithelial cell membranes specifically interacted with streptavidin-immobilized Fe3O4 magnetic nanoparticles and then formed inside-out-oriented cell membrane-coated magnetic nanoparticles (IOCMMNPs). The cell membrane orientation of the IOCMMNPs was successfully confirmed by immunogold electron microscopy, fluorescently labeled confocal microscopy, sialic acid quantification assay, and the adsorption capacity assay. Moreover, IOCMMNPs possessed satisfactory binding capacity, selectivity, and high sensitivity (limit of detection = 0.4 × 10-3 µg mL-1). Ultimately, IOCMMNPs successfully targeted two main compounds from Strychnos nux-vomica whose potential antitumor activities were further validated by pharmacological studies. The application of the inside-out cell membrane coating strategy further enhances the drug screening efficiency and broadens the insight and methodologies for drug lead discovery. This inside-out cell membrane coating concept also provides a method for the future development of engineered cell membrane-coated nanotechnology.

MSC-derived immunomodulatory extracellular matrix functionalized electrospun fibers for mitigating foreign-body reaction and tendon adhesion.

Adhesion formation during tendon healing remains a severe problem in clinical practice. Multiple factors contribute to postoperative adhesion formation, and macrophage-driven inflammation is thought to be greatly involved in this process. We hypothesize that reducing macrophage-mediated inflammation in the injured tendon by regulating M1 to M2 macrophage polarization may effectively inhibit adhesion formation. Here, we developed an acellular immunomodulatory biomaterial consisting of an electrospun polycaprolactone/silk fibroin (PCL/SF) composite fibrous scaffold functionalized with mesenchymal stem cell (MSC)-derived extracellular matrix (ECM). To enhance the immunoregulatory potential of MSCs, we performed inflammatory licensing with IFN-γ to obtain immunomodulatory ECM (iECM). Proteomic analyses of MSCs and their secreted ECM components from different culture conditions revealed the MSC-ECM molecular signatures and the potential mechanism of ECM immunoregulation. Then, the immunoregulatory potential of the iECM-modified scaffold was evaluated in vitro and in vivo. Relative to the PCL/SF fibrous scaffold, the iECM-functionalized scaffold facilitated M2 macrophage polarization and inhibited the expression of multiple cytokines (IL-1ß, IL-6, CXCL11, IL-10, IL-1R2, and TGF-ß1) in vitro, strongly suggesting the immunosuppressive ability of iECM derived from inflammatory licensed MSCs. Consistent with the in vitro findings, the results of rat subcutaneous implantation indicated that a markedly lower foreign-body reaction (FBR) was obtained in the PCL/SF-iECM group than in the other groups, as evidenced by thinner fibrotic capsule formation, less type I collagen production and more M2-type macrophage polarization. In the rat Achilles tendon injury model, the PCL/SF-iECM scaffold greatly mitigated tendon adhesion with clear sheath space formation between the tendon and the scaffold. These data highlight the immunomodulatory potential of iECM-functionalized fibrous scaffolds to attenuate FBR by modulating M2 macrophage polarization, thereby preventing tendon adhesion. STATEMENT OF SIGNIFICANCE: Electrospun PCL/SF fibrous scaffolds functionalized with ECM secreted by MSCs stimulated by inflammatory factor IFN-γ was developed that combined physical barrier and immunomodulatory functions to prevent tendon adhesion formation. PCL/SF micro-nanoscale bimodal fibrous scaffolds prepared by emulsion electrospinning possess high porosity and a large pore size beneficial for nutrient transport to promote intrinsic healing; moreover, surface modification with immunomodulatory ECM (iECM) mitigates the FBR of fibrous scaffolds to prevent tendon adhesion. The iECM-functionalized electrospun scaffolds exhibit powerful immunomodulatory potency in vitro and in vivo. Moreover, the iECM-modified scaffolds, as an anti-adhesion physical barrier with immunomodulatory ability, have an excellent performance in a rat Achilles tendon adhesion model. MSC secretome-based therapeutics, as an acellular regenerative medicine strategy, are expected to be applied to other inflammatory diseases due to its strong immunoregulatory potential.

Microcracks on the Rat Root Surface Induced by Orthodontic Force, Crack Extension Simulation, and Proteomics Study.

Root resorption is a common complication during orthodontic treatment. Microcracks occur on the root surface after an orthodontic force is applied and may be related to the root resorption caused by the orthodontic process. However, the mechanisms underlying root resorption induced by microcracks remain unclear. In this study, a rat orthodontic model was used to investigate the biological mechanisms of root resorption caused by microcracks. First, the first molar was loaded with 0.5-N orthodontic force for 7 days, and microcracks were observed on the root apex surface using a scanning electron microscope. Second, to describe the mechanical principle resulting in microcracks, a finite element model of rat orthodontics was established, which showed that a maximum stress on the root apex can cause microcrack extension. Third, after 7 days of loading in vivo, histological observation revealed that root resorption occurred in the stress concentration area and cementoclasts appeared in the resorption cavity. Finally, proteomics analysis of the root apex area, excluding the periodontal ligament, revealed that the NOX2, Aifm1, and MAPK signaling pathways were involved in the root resorption process. Microcrack extension on the root surface increases calcium ion concentrations, alters the proteins related to root resorption, and promotes cementoclast formation.

Molecular characterization of structural protein genes of dengue virus serotype 1 epidemic in Yunnan, Southwest China, in 2018.

A dengue virus serotype 1 (DENV-1) epidemic occurred from October to December 2018 in Xishuangbanna, Yunnan, Southwest China, neighboring Myanmar, Laos, and Vietnam. In this study, we investigated the molecular characteristics, evolution, and potential source of DENV from Xishuangbanna. The C (capsid), prM (premembrane), and E (envelope) genes of DENV isolated from 87 serum samples obtained from local patients were amplified and sequenced, and the sequences were evaluated by identification of mutations, phylogenetic and homologous recombination analysis, and secondary structure prediction. Phylogenetic analysis showed that all of the epidemic DENV strains from Xishuangbanna could be grouped in a branch with DENV-1 isolates, and were most similar to the Fujian 2005 (China, DQ193572) and Singapore 2016 (MF314188) strains. When compared with DENV-1SS (the standard strain), there were 31 non-synonymous mutations, but no obvious homologous recombination signal was found. Secondary structure prediction showed that some changes had occurred in a helical region in proteins of the MN123849 and MN123854 strains, but there were few changes in the disordered region. This study reveals the molecular characteristics of the structural genes of the Xishuangbanna epidemic strains in 2018 and provides a reference for molecular epidemiology, infection, and pathogenicity research and vaccine development.

CITCO as an Adjuvant Facilitates CHOP-Based Lymphoma Treatment in hCAR-Transgenic Mice.

Non-Hodgkin's lymphoma (NHL) is a malignant cancer originating in the lymphatic system with a 25-30% mortality rate. CHOP, consisting of cyclophosphamide (CPA), doxorubicin, vincristine, and prednisone, is a first-generation chemotherapy extensively used to treat NHL. However, poor survival rates among patients in advanced stages of NHL shows a need to improve this standard of care treatment. CPA, an integral component of CHOP, is a prodrug that requires CYP2B6-mediated bioactivation to 4-hydroxy-CPA (4-OH-CPA). The expression of CYP2B6 is transcriptionally regulated by the constitutive androstane receptor (CAR, NRi13). We have previously demonstrated that the induction of hepatic CYP2B6 by CITCO, a selective human CAR (hCAR) agonist, results in CHOP's enhanced antineoplastic effects in vitro. Here, we investigate the in vivo potential of CITCO as an adjuvant of CPA-based NHL treatment in a hCAR-transgenic mouse line. Our results demonstrate that the addition of CITCO to the CHOP regimen leads to significant suppression of the growth of EL-4 xenografts in hCAR-transgenic mice accompanied by reduced expression of cyclin-D1, ki67, Pcna, and increased caspase 3 fragmentation in tumor tissues. CITCO robustly induced the expression of cyp2b10 (murine ortholog of CYP2B6) through hCAR activation and increased plasma concentrations of 4-OH-CPA. Comparing to intraperitoneal injection, oral gavage of CITCO results in optimal hepatic cyp2b10 induction. Our in vivo studies have collectively uncovered CITCO as an effective facilitator for CPA-based NHL treatment with a pharmacokinetic profile favoring oral administration, promoting CITCO as a promising adjuvant candidate for CPA-based regimens.

Metformin Disrupts Bile Acid Efflux by Repressing Bile Salt Export Pump Expression.

PURPOSE: The bile salt export pump (BSEP), a key player in hepatic bile acid clearance, has been the center of research on drug-induced cholestasis. However, such studies focus primarily on the direct inhibition of BSEP, often overlooking the potential impact of transcriptional repression. This work aims to explore the disruption of bile acid efflux caused by drug-induced BSEP repression. METHODS: BSEP activity was analyzed in human primary hepatocytes (HPH) using a traditional biliary-clearance experiment and a modified efflux assay, which includes a 72-h pretreatment prior to efflux measurement. Relative mRNA and protein expressions were examined by RT-PCR and Western blotting, respectively. RESULTS: Metformin concentration-dependently repressed BSEP expression in HPH. Although metformin did not directly inhibit BSEP activity, longer metformin exposure reduced BSEP transport function in HPH by down-regulating BSEP expression. BSEP repression by metformin was found to be AMP-activated protein kinase-independent. Additional screening of 10 reported cholestatic non-BSEP inhibitors revealed that the anti-cancer drug tamoxifen also markedly repressed BSEP expression and reduced BSEP activity in HPH. CONCLUSIONS: Repression of BSEP alone is sufficient to disrupt hepatic bile acid efflux. Metformin and tamoxifen appear to be prototypes of a class of BSEP repressors that may cause drug-induced cholestasis through gene repression instead of direct BSEP inhibition.

Effect of Electrospun Silk Fibroin-Silk Sericin Films on Macrophage Polarization and Vascularization.

Biomaterial implantation is followed by an inflammatory cascade dominated by the macrophages, which polarized to the proinflammation M1 phenotype or prohealing M2 phenotype. Generally, silk sericin (SS) is considered to be of high immunogenicity associated with native silk fibers. The blends of silk fibroin (SF) and SS in different mass ratios might elicit different host immune responses and induce macrophage phenotype switch. The objective of this study was to assess the effects of electrospun SF-SS fibrous films with different mass ratios (10:0, 9:1, 8:2, and 7:3) on the macrophage phenotypes and explore the optimal ratio of SF and SS for angiogenesis. Our results indicated that the macrophages were activated by the addition of SS. When the mass ratio of SF and SS reached 7:3, the film displayed the highest degree of vascularization. The macrophages were induced to secrete more M1 and M2 cytokines accompanying with high M2/M1 ratio. Taken together, this study provided a perspective to promote neovascularization by modulating appropriate host response and macrophage phenotypes in tissue engineering field.

Simultaneous nano- and microscale structural control of injectable hydrogels via the assembly of nanofibrous protein microparticles for tissue regeneration.

Injectable hydrogels are advantageous as tissue regeneration scaffolds, as they can be delivered through a minimally invasive injection and seamlessly integrate with the target tissues. However, an important shortcoming of current injectable hydrogels is the lack of simultaneous control over their micro- and nanoscale structures. In this article, the authors report a strategy for developing injectable hydrogels that integrate a fibrous nanostructure and porous microstructure. The hydrogels are prepared by using novel nanofibrous microparticles as the building blocks. The protein based nanofibrous microparticles, fabricated by a spray freezing technology, can be injected through a syringe-needle system. A cell-compatible photocuring process can be deployed to connect the microparticles and form a mechanically robust hydrogel scaffold. The inter-particle voids combined to form the interconnected micropores and the diameter of the nanofibers (100-300 nm) closely mimics that of the native extracellular matrix. Compared to the non-porous hydrogels and non-fibrous hydrogels, the microparticle annealed nanofibrous (MANF) hydrogels potently enhance the osteogenic-marker expression (ALP, Runx2, OCT and BSP) and mineralization of human mesenchymal stem cells in vitro. MANF hydrogels also facilitate cell infiltration and enhance neovasculization in a subcutaneous implantation model in vivo. The capacity of MANF hydrogels to promote bone regeneration is investigated in a calvarial bone repair model. MANF hydrogels demonstrate significant higher bone regeneration after 8 weeks, indicating the significant role of microporosity and nanofibrous architecture in bone regeneration.

Metazachlor: Mode of action analysis for rat liver tumour formation and human relevance.

In a 2-year study the herbicide metazachlor (BAS 479H) was shown to significantly increase the incidence of liver tumours in female Wistar rats at a dietary level of 8000 ppm. As metazachlor is not a genotoxic agent, a series of in vivo and in vitro investigative studies were undertaken to elucidate the mode of action (MOA) for metazachlor-induced female rat liver tumour formation. Male and female Wistar rats were given diets containing 0 (control), 200 and 8000 ppm metazachlor for 3, 7, 14 and 28 days. The treatment of male rats with 200 and 8000 ppm metazachlor and female rats with 8000 ppm metazachlor resulted in significant increases in relative liver weight, which was associated with a centrilobular hepatocyte hypertrophy. Hepatocyte replicative DNA synthesis (RDS) was significantly increased in male rats given 8000 ppm metazachlor for 3 and 7 days and in female rats given 200 ppm metazachlor for 7-28 days and 8000 ppm metazachlor for 3-28 days. Significant increases in relative liver weight, centrilobular hepatocyte hypertrophy and hepatocyte RDS were also observed in male and female Wistar rats given and 500 ppm sodium phenobarbital (NaPB) for 3-28 days. The treatment of female Wistar rats with either 8000 ppm metazachlor for 7 days or with 500 ppm NaPB for 3 and 7 days resulted in the nuclear translocation of the hepatic constitutive androstane receptor (CAR). Treatment of male and female Wistar rats with 8000 ppm metazachlor for 14 days resulted in significant increases in hepatic microsomal total cytochrome P450 (CYP) content, CYP2B subfamily-dependent enzyme activities and mRNA levels, together with some increases in CYP3A enzyme activity and mRNA levels. The treatment of male Wistar rat hepatocytes with metazachlor (concentration range 0.5-50 µM) and NaPB (500 µM) for 4 days resulted in increased CYP2B enzyme activities and mRNA levels; with metazachlor and NaPB also producing significant increases in hepatocyte RDS levels. Studies were also performed with hepatocytes from male Sprague-Dawley wild type (WT) rats and CAR knockout (CAR KO) rats. While both treatment with metazachlor and NaPB for 4 days increased CYP2B enzyme activities and mRNA levels in WT rat hepatocytes, only minor effects were observed in CAR KO rat hepatocytes. Treatment with both metazachlor and NaPB only increased RDS in WT but not in CAR KO rat hepatocytes. The treatment of hepatocytes from two male human donors with 0.5-25 µM metazachlor or 500 µM NaPB for 4 days resulted in increases in CYP2B6 and CYP3A4 mRNA levels but had no effect on hepatocyte RDS. EGF as concurrently used positive control demonstrated the expected RDS response in all rat and human hepatocyte cultures. In conclusion, a series of in vivo and in vitro investigative studies have demonstrated that metazachlor is a CAR activator in rat liver, with similar properties to the prototypical CAR activator phenobarbital. A robust MOA for metazachlor-induced female rat liver tumour formation has been established. Based on the lack of effect of metazachlor on RDS in human hepatocytes, it is considered that the MOA for metazachlor-induced rat liver tumour formation is qualitatively not plausible for humans.