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Background Tumor-associated macrophages (TAMs) are the most abundant stromal cells in the tumor microenvironment. Turning the TAMs against their host tumor cells is an intriguing therapeutic strategy particularly attractive for patients with immunologically “cold” tumors. This concept was mechanistically demonstrated on in vitro human and murine lung cancer cells and their corresponding TAM models through combinatorial use of nanodiamond-doxorubicin conjugates (Nano-DOX) and a PD-L1 blocking agent BMS-1. Nano-DOX are an agent previously proved to be able to stimulate tumor cells’ immunogenicity and thereby reactivate the TAMs into the anti-tumor M1 phenotype. Results Nano-DOX were first shown to stimulate the tumor cells and the TAMs to release the cytokine HMGB1 which, regardless of its source, acted through the RAGE/NF-κB pathway to induce PD-L1 in the tumor cells and PD-L1/PD-1 in the TAMs. Interestingly, Nano-DOX also induced NF-κB-dependent RAGE expression in the tumor cells and thus reinforced HMGB1’s action thereon. Then, BMS-1 was shown to enhance Nano-DOX-stimulated M1-type activation of TAMs both by blocking Nano-DOX-induced PD-L1 in the TAMs and by blocking tumor cell PD-L1 ligation with TAM PD-1. The TAMs with enhanced M1-type repolarization both killed the tumor cells and suppressed their growth. BMS-1 could also potentiate Nano-DOX’s action to suppress tumor cell growth via blocking of Nano-DOX-induced PD-L1 therein. Finally, Nano-DOX and BMS-1 achieved synergistic therapeutic efficacy against in vivo tumor grafts in a TAM-dependent manner. Conclusions PD-L1/PD-1 upregulation mediated by autocrine and paracrine activation of the HMGB1/RAGE/NF-κB signaling is a key response of lung cancer cells and their TAMs to stress, which can be induced by Nano-DOX. Blockade of Nano-DOX-induced PD-L1, both in the cancer cells and the TAMs, achieves enhanced activation of TAM-mediated anti-tumor response. Graphic abstract

Background The side effects of chemotherapy-induced nausea and vomiting (CINV) and myelosuppression reduce the cancer patients’ adherence to chemotherapy. Many Chinese patients choose Chinese medicine (CM) during chemotherapy to reduce side effects; however, the evidence is lacking. The efficacy of a CM herbal treatment protocol, Jianpi Bushen Sequential Formula (健脾补肾续贯方, JBSF) will be evaluated on chemotherapy completion rate among patients with colon cancer. Methods A multi-center double-blind randomized controlled trial (RCT) will be conducted on 400 patients with colon cancer who will receive 8 cycles of adjuvant chemotherapy with oxaliplatin and capecitabine (CAPEOX). Patients will be randomized 1:1 to receive the JBSF or placebo formula. The primary outcome is the overall chemotherapy completion rate. The secondary outcomes include individual chemotherapy completion rate, 4-cycle completion rate of chemotherapy, time to treatment failure, relative dose intensity and treatment toxicity. Follow-up visits will be scheduled before every and after last chemotherapy. Discussion This study will provide evidence on whether JBSF can improve the chemotherapy completion rate and reduce side effects among patients with colon cancer. (Trial registration: ClinicalTrials.gov, No. NCT03716518)

Chlorin e6 (Ce6) is a promising photosensitizer for tumor photodynamic therapy (PDT). However, the efficacy of Ce6 PDT is limited by Ce6's poor water solubility, rapid blood clearance, and inadequate accumulation in the tumor tissue. This problem is tackled in this work, wherein functionalized superparamagnetic iron oxide nanoparticles (IO-NPs) were used as carriers to deliver Ce6 to melanoma. The IO-NPs were coated with polyglycerol (PG) to afford good aqueous solubility. The chemotherapeutic agent doxorubicin (DOX) was attached to the PG coating via the hydrazone bond to afford affinity to the cell membrane and thereby promote the cell uptake. The hydrophobic nature of DOX also induced the aggregation of IO-NPs to form nanoclusters. Ce6 was then loaded onto the IO nanoclusters through physical adsorption and coordination with surface iron atoms, yielding the final composites IO-PG-DOX-Ce6. In vitro experiments showed that IO-PG-DOX-Ce6 markedly increased Ce6 uptake in mouse melanoma cells, leading to much-enhanced photocytotoxicity characterized by intensified reactive oxygen species production, loss of viability, DNA damage, and stimulation of tumor cell immunogenicity. In vivo experiments corroborated the in vitro findings and demonstrated prolonged blood clearance of IO-PG-DOX-Ce6. Importantly, IO-PG-DOX-Ce6 markedly increased the Ce6 distribution and retention in mouse subcutaneous melanoma grafts and significantly improved the efficacy of Ce6-mediated PDT. No apparent vital organ damage was observed at the same time. In conclusion, the IO-PG-DOX NPs provide a simple and safe delivery platform for efficient tumor enrichment of Ce6, thereby enhancing antimelanoma PDT.

Astrocytes are key stromal components in glioblastoma (GBM) and have complex interactions with the GBM cells (GBC) promoting the survival, progression and therapy resistance of GBM. In this study, we first demonstrated the existence of a reciprocal activation loop mediated by the STAT3/IL-6 signaling between GBC and astrocytes. This loop of reciprocity was found to be initiated by the constitutive activity of STAT3 and downstream expression of IL-6 in the GBC. GBC-derived IL-6 activated STAT3 and thereby upregulated IL-6 expression in the astrocytes. Astrocyte-derived IL-6 acted back on the GBC causing further activation of STAT3 and leading to enhanced downstream events that promote proliferation, migration, invasion and apoptosis resistance of the GBC. Next, we showed that doxorubicin-polyglycerol-nanodiamond conjugates (Nano-DOX), which could be delivered via GBM-associated macrophages, suppressed STAT3 activity in the GBC reducing their IL-6 output to the astrocytes and thereby abolished the astrocytes' feedback activation of the GBC. Moreover, Nano-DOX also suppressed stimulated activation of STAT3 and IL-6 induced by temozolomide, a first-line anti-GBM chemotherapy, resistance to which critically involves STAT3 activation. In conclusion, Nano-DOX could disrupt the STAT3/IL-6-mediated reciprocal activation loop between the GBC and astrocytes. Nano-DOX also provides a novel approach to therapeutic modulation of the GBM microenvironment.

A new lignanamide derivative, lycichinenamide (1), has been isolated from the root barks of Lycium chinense together with nine known compounds, lyciumamide H (2), 5,2′-dihydroxy-6,7,8,6′-tetramethoxyflavone (3), 7α,8β-3′-O-methylcedrusin (4), smiglabrol (5), trans-N-caffeoyltyramine (6), vanillic acid (7), syringic acid (8), 2,6-dimethoxy-1,4-benzoquinone (9), and 1,5-dihydroxy-2-isoprenyl-3-methoxyxanthone (10). The structure of the new compound 1 was determined through spectroscopic and MS analyses. Among the isolates, lycichinenamide (1), lyciumamide H (2), smiglabrol (5), and trans-N-caffeoyltyramine (6) showed potent inhibition with IC50 values of 18.5 ± 2.1, 21.8 ± 1.8, 12.4 ± 0.3, and 24.6 ± 2.0 μM, respectively, against LPS-induced NO generation. In addition, 1,5-dihydroxy-2-isoprenyl-3-methoxyxanthone (10) exhibited anti-Dengue virus activity with IC50 value of 22.4 ± 1.6 μM against Dengue virus serotype 2 strain PL046.

Purpose To assess the effectiveness and safety of acupuncture for erectile dysfunction (ED). Materials and methods We searched six major English and Chinese databases included randomized controlled trials (RCTs) testing acupuncture alone or in combination for ED. Dichotomous data were presented as risk ratio (RR) and continuous data were presented as mean difference (MD) both with 95% confidence interval (CI). The Revman (v.5.3) was used for data analyses. Quality of evidence across studies was assessed by the online GRADEpro tool. Results We identified 22 RCTs, fourteen of them involving psychogenic ED. Most of the included RCTs had high or unclear risk of bias. There was no difference between electro-acupuncture and sham acupuncture with electrical stimulation on the rate of satisfaction and self-assessment (RR, 1.50; 95% CI, 0.71-3.16; 1 trial). Acupuncture combined with tadalafil appeared to have better effect on increasing cure rate (RR, 1.31; 95% CI, 1.00-1.71; 2 trials), and International Index of Erectile Function-5 scores (MD, 5.38; 95% CI, 4.46-6.29; 2 trials). When acupuncture plus herbal medicine compared with herbal medicine alone, the combination therapy showed significant better improvement in erectile function (RR, 1.68; 95% CI, 1.31-2.15; 7 trials). Only two trials reported facial red and dizziness cases, and needle sticking and pruritus cases in acupuncture group. Conclusions Low quality evidence shows beneficial effect of acupuncture as adjunctive treatment for people mainly with psychogenic ED. Safety of acupuncture was insufficiently reported. The findings should be confirmed in large, rigorously designed and well-reported trials.

Photodynamic therapy (PDT) is an emerging anti-tumor strategy. Photosensitizer chlorin e6 (Ce6) can induce photodynamic effect to selectively damage lung cancer cells. In order to further improve its tumor targeting ability, macrophages can be applied as carrier to deliver Ce6 to lung cancer. Tumor associated macrophages (TAM) are important immunocytes in lung cancer immune microenvironment. TAM play crucial role in tumor promotion due to the Immunosuppressive property, reprogramming phenotype of TAM therefore has become a promising strategy. Based on this, in the present study, we suppose that TAM can be used as carrier to deliver Ce6 to lung cancer and be reprogrammed to M1 phenotype by photodynamic action to mediate anti-lung cancer efficacy. The results showed TAM could load with Ce6 and keep viability in the absence of near infrared irradiation (NIR). Moreover, Its viability decreased little within 10 h after NIR. Ce6-loaded TAM could deliver Ce6 to lung cancer cells and retain some drugs in TAM per se. After NIR, phagocytosis of macrophages was enhanced. The expressions of GBP5, iNOS and MHC-II was up-regulated, which indicated TAM were polarized to M1 phenotype. Finally, the study also found the reprogrammed macrophages could inhibit the proliferation and promote the apoptosis of lung cancer cells. These results suggested that macrophages could deliver Ce6 to lung cancer and exhibit anti-lung cancer effect through photodynamic reprogramming. This study provides a novel approach for combining photodynamic action with anti-tumor immunotherapy.

With the increased interest in the practical use of hydrochar, concerns about the possible environmental biotoxicity of hydrochar and its released dissolved organic matters (DOM) have grown. As a common method for removing bio-oil on the surface of hydrochar, the effect of organic solvent washing on the properties of hydrochar released DOM remains unclear. In this study, we made a comprehensive comparison of hydrochar properties and molecule structure as well as biotoxicity of DOM released from HC (raw hydrochar) and THC (hydrochar washed by tetrahydrofuran). The results indicated that the mass loss of hydrochar was obvious after tetrahydrofuran (THF) washing, and a decline of H/C atomic ratio and increase of N/C and O/C atomic ratios was observed based on Van Krevelen (VK) diagram. This result was further confirmed by FTIR, 13C NMR, and XPS results. Meanwhile, the molecule structure of DOM was shifted to lower molecule weight with higher O-contain compounds after THF extraction due to the demethanation process. However, the biotoxicity experiments indicated that both extracted DOM had no significant impact on germination rate of wheat, and HC-treated sample even exhibited growth superiority. Nevertheless, potential toxicity was observed with the increase of the activity of antioxidant enzymes, and THF washing aggravated the potential oxidative damage through increasing the aromaticity of DOM. Such understanding highlights the importance of evaluating hydrochar and its released DOM before applications, so as to reduce the potential environment biotoxicity.

Immunosuppression is a salient feature of GBM associated with the disease's grim prognosis and the limited success of anti-GBM immunotherapy. Stimulating immunogenicity of the GBM cells (GC) is a promising approach to subverting the GBM-associated immunosuppression. We had previously devised a drug composite based on polyglycerol-functionalized nanodiamonds bearing doxorubicin (Nano-DOX) and demonstrated that Nano-DOX effectively modulated GBM's immunosuppressive microenvironment through stimulating the immunogenicity of GC and initiated anti-GBM immune responses. The present study now explored the mechanism of Nano-DOX's immunostimulatory action. Nano-DOX was found to induce autophagy rather than apoptosis in GC and stimulated GC to emit antigens and damage-associated molecular patterns (DAMPs) that are potent adjuvants, which resulted in enhanced activation of dendritic cells (DC). Heightened autophagosome release was observed in Nano-DOX-treated GC but was shown not to be a major channel of antigen donation. Blocking autophagy in GC not only reduced Nano-DOX-stimulated GC antigen donation and DAMPs emission, but also efficiently attenuated DC activation stimulated by Nano-DOX-treated GC. Taken together, these findings suggest that activation of autophagy is a central mechanism whereby Nano-DOX stimulates GC's immunogenicity. Our work provides new insight on how nanotechnology can be applied to therapeutically modulate the GBM immune microenvironment by harnessing autophagy in the cancer cells. STATEMENT OF SIGNIFICANCE: Immunosuppression is a salient feature of GBM associated with the grim prognosis of the disease and the limited success of anti-GBM immunotherapy. We demonstrated that Doxorubicin-polyglycerol-nanodiamond composites could activate autophagy in GBM cells and thereby stimulate the immunogenecity of GBM cells. This discovery 1, sheds new light on how nanotechnology could be applied to therapeutically modulate the tumor immune microenvironment, and 2, provides a powerful tool for subverting the GBM's immunosuppressive microenvironment, which has great therapeutic potential for the treatment of GBM.

Background Laparoscopic hepatectomy (LH) is growing in popularity, but its efficacy and safety are still controversial. Few multicenter, large, population-based, prospective, randomized studies have compared LH with open hepatectomy (OH). We performed a meta-analysis to compare the treatment outcome of patients undergoing LH versus OH. Methods We searched PubMed, EMBASE, Cochrane Central Register, CNKI, and WanFang Med Online databases to November 30th, 2016 for randomized controlled trials (RCTs) that compared treatment outcome for LH and OH. Eligibility criteria included evaluation of operation time, blood loss, complications, and hospital stay after surgery for adult patients who underwent LH or OH. Reviewers in pairs independently screened the studies, extracted data and assessed the risk of bias of included studies. Agreement was achieved. RevMan 5.3 was used to conduct meta-analysis. Complete case analysis was used as primary analysis. Predefined subgroup analysis includes benign and malignant disease. Results Eight RCTs with a total of 554 patients were included in the meta-analysis, 275 types of LH and 279 types of OH. LH reduced the hospital stay after surgery (8 trials, 554 patients, MD = −3.84 days, 95%CI: −5.05 to −2.63, P Conclusions Laparoscopy is more effective in terms of hospital stay after surgery and time to first flatus with fewer blood loss and complication rate for hepatic resection compared with open surgery.

OBJECTIVE Photodynamic therapy (PDT) has emerged as a promising strategy in the treatment of malignant tumors due to its high selectivity, non-toxicity, and non-invasiveness. However, PDT can also induce DNA damage and subsequent repair response, which may reduce the efficacy of PDT. In the present study, we sought to explore the effect of chlorin e6 (Ce6)-mediated PDT on DNA damage and DNA damage response (DDR) in lung cancer cells. In addition, the effect of PDT combined with ATM inhibitor on molecules of DDR and the possibility of improving the efficacy of PDT were further investigated. MATERIALS AND METHODS In the in vitro study, lewis cells were submitted to Ce6 treatment (2, 4, 8, 16, 32 μg/mL). To determine the concentration of Ce6, uptake and toxicity of Ce6 mediated PDT were detected using flow cytometry (FACS), Confocal microscopy, and CCK-8. In the subsequent research, 8 μg/mL of Ce6 was the treatment condition for inducing PDT. The different post-irradiation placement times were further grouped under this condition (2, 4, 6, 12 h). Cellular reactive oxygen species (ROS), damage of DNA were measured by DCFH-DA probe, comet assay respectively. Then the expression of p-ATM, p53, and γ-H2A.X proteins related to DNA damage response, was detected by WB. The efficacy of Ce6 induced PDT was also demonstrated by Annexin-V/PI staining as well as the expression of PCNA, cleaved-caspase-3. On this basis, ATM inhibitor was applied to treat lewis cells combined with Ce6 (2, 4 h) to investigate whether the efficacy of PDT induced by Ce6 can be improved after the ATM-related DDR was blocked. The cell viability, apoptosis, and expression of associated proteins were assayed. RESULTS At 2-4 h after PDT treatment, ROS was dramatically elevated in lewis cells, DNA double-strand breaks (DDSB) occurred, as well as up-regulation of DDR proteins γ-H2A.X, p-ATM, and p53. At the same time, lewis cells did not undergo significant apoptosis. After ATM inhibition, the DDR was significantly blocked within 2-4 hours after Ce6 induced PDT, along with a pronounced decrease in cell viability followed by a prominent increase of apoptosis. CONCLUSION Ce6-mediated PDT generates ROS in a short period time, thus inducing DNA damage, ATM-related DDR as well as promoting resistance of lung cancer cells to PDT. Combining ATM inhibitor with PDT could effectively inhibit the DDR induced by PDT, thereby enhancing the efficacy. This study reveals a new resistance mechanism of PDT and proposes an intervention strategy.

Aim: To mechanistically prove the concept of monocyte-mediated nano drug delivery in glioblastoma (GBM). Results: nano-doxorubicin-loaded monocytes (Nano-DOX-MC) were viable, able to cross an artificial endothelial barrier and capable of infiltrating GBM spheroids and releasing drug therein. GBM cells stimulated unloading of Nano-DOX-MC and took up the unloaded drug and released damage-associated molecular patterns. In mice with orthotopic GBM xenografts, Nano-DOX-MC resulted in much improved tumor drug delivery efficacy and damage-associated molecular patterns emission. Mechanistically, Nano-DOX was found sequestered in the lysosomal compartment and to induce autophagy, which may underlie MC's tolerance to Nano-DOX. Lysosomal exocytosis was found involved in the discharging mechanism of intracellular Nano-DOX. Conclusion: Nano-DOX can be effectively delivered by MC in GBM and induce cancer cell damage.

Background: Peri-procedural myocardial injury (PMI) during percutaneous coronary intervention (PCI) will result in an unfavorable clinical prognosis in patients, thus urgently necessitating effective drug treatment measures. Shen-Yuan-Dan (SYD) capsules are a traditional Chinese medicine (TCM) preparation that have been found to have potential myocardial protection effects during the peri-procedural phase of PCI in previous clinical and basic research; however, there is a lack of rigorous, randomized, and controlled studies. The aim of this study is to evaluate the efficacy and safety of SYD in decreasing PMI. Methods: This is a randomized, double-blind, placebo-controlled clinical trial. A total of 284 patients with unstable angina will be randomized into test and control groups. The two groups will be given SYD or a placebo (three times each day, four capsules each time) 3 days before PCI on the basis of conventional treatment. Twelve hours before PCI, an additional 4 capsules will be given, and drug treatment is planned to be maintained for 1 month after surgery. Dynamic changes in the myocardial enzyme in four time-points (before PCI, and 4, 24, 48 hours after PCI) in both groups of patients that will be observed. The follow-up period will be 1 month. The primary observation markers are planned to evaluate the efficacy and safety of SYD in decreasing PMI. The secondary observation markers will be to evaluate the major adverse cardiovascular events (MACEs) status at day 30 after PCI, (all-cause mortality, non-fatal myocardial infarction, repeated revascularization of target blood vessel) and Seattle Angina Questionnaire scores. GRACE scores will be used for risk stratification, and the intervention efficacy of SYD on PMI patients with different risks will be retrospectively evaluated. Discussion: This study will provide a rigorous clinical evidence to evaluate the efficacy and safety of SYD in decreasing PMI and the results are worth anticipating. Trial registration: The design of this trial has been registered with the Chinese Clinical Trial Registry (No. ChiCTR-IPR-17011069).

Aim: To mechanistically compare the effects of doxorubicin (DOX) and DOX conjugated with nanodiamonds (Nano–DOX) on human glioblastoma cells (GC). Materials & methods: GC viablity, proliferation and activation of apoptosis and autophagy was assayed in response to DOX and Nano–DOX. Expression and release of HMGB1 were measured and its role in apoptosis and autophagy probed in response to DOX and Nano–DOX. Results: DOX induced apoptosis in GC while Nano–DOX induced autophagy. Inhibition of autophagy in Nano–DOX-treated GC promoted apoptosis. DOX suppressed the emission of HMGB1 while Nano–DOX stimulated HMGB1 emission which was attenuated when autophagy was repressed. Blocking of HMGB1 emission mitigated autophagy and enhanced apoptosis in Nano–DOX-treated GC. Exogenously administered HMGB1 promoted autophagy and protected against apoptosis in both Nano–DOX-treated GC and DOX-treated GC. Conclusions: Nano–DOX is a potent autophagy activator as opposed to DOX as an apoptosis inducer. Nano–DOX initiates a mutual reinforcement loop between autophagy and HMGB1 in GC and thereby protects GC against apoptosis.

Glioblastoma (GBM) is the deadliest and most common type of primary brain tumor in adults with a grim prognosis despite multimodal treatments. Dendritic cell (DC)-based immunotherapy has emerged as a promising therapeutic modality for GBM, whose efficacy is nonetheless fundamentally undermined by GBM-induced immunosuppression. Inducing emission of damage associated molecular patterns (DAMPs) is a highly effective strategy to subvert tumor-associated immunosuppression. The present work was carried out to explore the idea of subverting the GBM immunosuppressive microenvironment through DC-mediated delivery of doxorubicin-polyglycerol-nanodiamond composites (Nano-DOX), a potent DAMPs inducer demonstrated by our previous study, and thereby eliciting enhanced DC-driven anti-GBM immune response. In the in-vitro work on human cell models, Nano-DOX-loaded DC were shown to be functionally viable and release cargo drug to co-cultured GBM cells (GC). Nano-DOX-treated GC displayed not only profuse DAMPs emission but also antigen release. Enhanced activation and acquisition and presentation of GC-derived antigen were then demonstrated in DC in co-culture with GC and Nano-DOX. Consistently, co-culture with GC and Nano-DOX also activated mouse bone marrow-derived DC (mDC) which in turn stimulated mouse spleen-derived lymphocytes which ultimately suppressed co-cultured GC. Next, athymic mice bearing orthotopic human GBM xenografts were intravenously injected with Nano-DOX-loaded mDC and, 48 h later, spleen-derived lymphocytes. The presence of Nano-DOX, DAMPs emission and enhanced infiltration and activation of mDC and lymphocytes were detected in the GBM xenografts. Taken together, our results demonstrate the efficacy of DC-mediated delivery of Nano-DOX to stimulate GC immunogenicity and elicit anti-cancer immune response in the GBM. By this work, we present a novel approach with great application potential to subverting the GBM immunosuppressive microenvironment and to anti-GBM immunotherapy. Investigation has also been conducted probing the mechanisms by which Nano-DOX stimulates GC immunogenicity, which is described in a follow-up paper.

Summary In the developing brain, immature synapses contain calcium-permeable AMPA glutamate receptors (AMPARs) that are subsequently replaced with GluA2-containing calcium-impermeable AMPARs as synapses stabilize and mature. Here, we show that this essential switch in AMPARs and neuronal synapse maturation is regulated by astrocytes. Using biochemical fractionation of astrocyte-secreted proteins and mass spectrometry, we identified that astrocyte-secreted chordin-like 1 (Chrdl1) is necessary and sufficient to induce mature GluA2-containing synapses to form. This function of Chrdl1 is independent of its role as an antagonist of bone morphogenetic proteins (BMPs). Chrdl1 expression is restricted to cortical astrocytes in vivo, peaking at the time of the AMPAR switch. Chrdl1 knockout (KO) mice display reduced synaptic GluA2 AMPARs, altered kinetics of synaptic events, and enhanced remodeling in an in vivo plasticity assay. Studies have shown that humans with mutations in Chrdl1 display enhanced learning. Thus astrocytes, via the release of Chrdl1, promote GluA2-dependent synapse maturation and thereby limit synaptic plasticity.

Glioblastoma (GBM) is the most frequent and malignant brain tumor with a high mortality rate. The presence of a large population of macrophages (Mφ) in the tumor microenvironment is a prominent feature of GBM and these so-called tumor-associated Mφ (TAM) closely interact with the GBM cells to promote the survival, progression and therapy resistance of the GBM. Various therapeutic strategies have been devised either targeting the GBM cells or the TAM but few have addressed the cross-talks between the two cell populations. The present study was carried out to explore the possibility of exploiting the cross-talks between the GBM cells (GC) and TAM for modulation of the GBM microenvironment through using Nano-DOX, a drug composite based on nanodiamonds bearing doxorubicin. In the in vitro work on human cell models, Nano-DOX-loaded TAM were first shown to be viable and able to infiltrate three-dimensional GC spheroids and release cargo drug therein. GC were then demonstrated to encourage Nano-DOX-loaded TAM to unload Nano-DOX back into GC which consequently emitted damage-associated molecular patterns (DAMPs) that are powerful immunostimulatory agents as well as indicators of cell damage. Nano-DOX was next proven to be a more potent inducer of GC DAMPs emission than doxorubicin. As a result, Nano-DOX-damaged GC exhibited an enhanced ability to attract both TAM and Nano-DOX-loaded TAM. Most remarkably, Nano-DOX-damaged GC reprogrammed the TAM from a pro-GBM phenotype to an anti-GBM phenotype that suppressed GC growth. Finally, the in vivo relevance of the in vitro findings was tested in animal study. Mice bearing orthotopic human GBM xenografts were intravenously injected with Nano-DOX-loaded mouse TAM which were found releasing drug in the GBM xenografts 24 h after injection. GC damage was evidenced by the induction of DAMPs emission within the xenografts and a shift of TAM phenotype was detected as well. Taken together, our results demonstrate a novel way with therapeutic potential to harness the cross-talk between GBM cells and TAM for modulation of the tumor immune microenvironment.AbbreviationsATP, adenosine triphosphate; BBB, blood-brain barrier; BCA, bicinchoninic acid; BMDM, bone marrow derived macrophages; CD, cluster of differentiation; CFSE, 5(6)-carboxyfluorescein diacetate, succinimidyl ester; CM, conditioned culture medium; CNS, central nervous system; CRT, calreticulin; DAMPs, damage-associated molecular patterns; DAB, diaminobenzidine; DOX, doxorubicin; ECL, enhanced chemiluminescence; ELISA, enzyme-linked immunosorbent assay; HMGB1, high mobility group protein B1; HSP90, heat shock protein 90; FACS, flow cytometry; GBM, glioblastoma; Guanylate Binding Protein 5 (GBP5); GC, glioblastoma cells; IHC, immunohistochemical; IL, interleukin; Mφ, macrophages; mBMDM, mouse BMDM; mBMDM2, Type-2 mBMDM; M1, Type-1 Mø; M2, Type-2 Mø; Nano-DOX, ND-PG-RGD-DOX; ND, nanodiamonds; Nano-DOX-mBMDM, Nano-DOX-loaded mouse BMDM; NGCM, Nano-DOX-treated-GC-conditioned medium; PBS, phosphate buffered saline; PG, polyglycerol; PMA, phorbol 12-myristate 13-acetate; PVDF, polyvinylidene fluoride; RGD, tripeptide of L-arginine, glycine and L-aspartic acid; RM, regular culture medium; SD, standard deviation; TAM, tumor-associated Mφ; TBST, Tris Buffered Saline with Tween® 20.Graphic abstract

Objective: The 2019 novel coronavirus (2019-nCoV) outbreak has escalated into a global pandemic. According to Chinese guidance for coronavirus disease-19 (COVID-19): Prevention, control, diagnosis, and management, Reduning injection can effectively treat, the disease caused by the virus. To identify the active ingredients of Reduning injection and COVID-19 disease-related pathways, we conducted a network pharmacology study. Methods: The Traditional Chinese Medicine Systems Pharmacology database was used to screen the chemical constituents and potential targets of Reduning injection. The gene names were converted to the corresponding protein names using UniProt. GeneCards and OMIM databases were used to select targets related to 2019-nCoV. Using Cytoscape 3.7.2 software platform and STRING database, we constructed drug-common target and target protein protein-protein interaction network diagrams. Rx64 3.6.2 software and Bioconductor biological information software package were used for Gene Ontology (GO) functional enrichment and KEGG pathway analyses. Results: In Reduning injection, a total of 33 effective chemical components were obtained that were involved in 151 signaling pathways, of which 44 targets were considered therapeutically relevant. Conclusion: Reduning injection can be potentially applied for the treatment of COVID-19 based on the results of our network pharmacology study.

To study the clinical, biochemical, and genetic heterogeneity of six Chinese patients and their mothers with the 3243 AG mutation, six patients (ranging from 5 to 11 years) were hospitalized. All the mothers were healthy. Mitochondrial respiratory chain enzyme activities were determined by spectrophotometry. Mitochondrial gene was analyzed in all patients. Six core pedigrees were investigated. Two patients had mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes syndrome and one had Leigh syndrome. The common initial symptoms were headache, vomiting, blurred vision, and epilepsy. m.3243AG mutation was detected in all patients and their mothers. The mutation loads ranged from 43.6% to 58% and those of their mothers ranged from 14.1% to 28.6%. Varied respiratory chain deficiencies were observed in all patients and two mothers. m.3243AG mutation can result in a wide spectrum of respiratory chain complex deficiencies. Mitochondrial DNA mutation detected in blood may be likely to transmit to offspring, and the mutation load may increase.

Mitochondrial respiratory chain complex I enzyme deficiency is the most commonly seen mitochondrial respiratory chain disorder. Although screening and diagnostic methods are available overseas, clinically feasible diagnostic methods have not yet been established in China. In this study, four Chinese boys with Leigh syndrome due to complex I deficiency were diagnosed by mitochondrial respiratory chain enzyme assay and DNA analysis using peripheral blood leukocytes. Four patients were admitted at the age of 5-14 years because of unexplained progressive neuromuscular symptoms, including motor developmental delay or regression, weakness, and seizures. Their cranial magnetic resonance imaging revealed typical finding as Leigh syndrome. Peripheral leukocyte mitochondrial respiratory chain complex I activities were found decreased to 9.6-33.1 nmol/min/mg mitochondrial protein(control 44.0 ± 5.4 nmol/min/mg). The ratios of complex I to citrate synthase activity were also decreased (8.9-19.8% in patients vs. control 48 ± 11%). Three mtDNA mutations were identified from three out of four patients, supporting the diagnosis of complex I deficiency. Point mutations m.10191TC in mitochondrial ND3 gene, m.13513GA in ND5 gene and m.14,453GA in ND6 gene were detected in three patients.

Mitochondrial oxidative phosphorylation defects are a common cause of mitochondrial diseases, which are characterized by multiorgan involvement and clinically heterogeneous manifestations. Diagnosis is difficult because of the lack of clinically feasible methods. In this study, mitochondrial complex I-V enzyme activity was measured in 64 patients with suspected mitochondrial disease and 200 healthy controls. Spectrophotometric assay was used for the analysis of mitochondrial complex I-V enzyme activity in peripheral leukocytes. Diagnosis was based on clinical presentation, magnetic resonance imaging (MRI), muscle pathology, and point mutation screening in mitochondrial DNA. The differential diagnosis of aminoacidopathies, organic acidurias, and fatty acid β-oxidation defects was made. Thirty-five patients (54.7%) were diagnosed with Leigh syndrome based on characteristic brain MRI. Complex enzyme activity in controls was found to be stable. A deficiency in the oxidative phosphorylation was found in 29 patients (45.3%). Twenty (31.2%) patients had isolated complex defects, complex I deficiency (n = 2, 3.1%), complex II deficiency (n = 3, 4.7%), complex III deficiency (n = 5, 7.8%), complex IV deficiency (n = 5, 7.8%), and complex V deficiency (n = 5, 7.8%). Nine patients were found to have combined deficiencies, 3 (4.7%) had combined deficiencies of complex I and IV, 2 (3.1%) had combined deficiencies of complex III and V, and 2 (3.1%) had a combined deficiency of complex I and V. In conclusion, the peripheral leukocyte oxidative phosphorylation enzyme activity assay was found to be a reliable method for the diagnosis of mitochondrial diseases.

Many traditional Chinese medicines have been used as immunomodulators that act as either immunosuppressants or immunostimulators. Recently, our lab successfully isolated a monomer telocinobufagin (TCB) from the chloroform extract of Chan Su (Venenum Bufonis). In the present paper, we evaluated the immunomodulatory effects of this compound in vitro. We found that TCB significantly stimulates splenocyte proliferation when administered alone or in combination with polyclonal T-cell mitogens concanavalin A (Con A) and lipopolysaccharide. Telocinobufagin markedly enhances natural killer cell and peritoneal macrophage activation. Telocinobufagin increases the percentage of CD4, CD8 positive cells within a population of splenocytes. Moreover, we found that the level of several Th1 cytokines, including interleukin-2 (IL-2), interleukin-12 (IL-12), interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha), are significantly increased after TCB treatment, while the level of the Th2 cytokine interleukin-4 (IL-4) is significantly decreased. As a result, the ratio of Th1/Th2 is significantly increased. Taken together, these results indicate that TCB has potential immune system regulatory effects and suggest that this compound could be developed as a novel immunotherapeutic agent to treat cancer and other immune-mediated diseases, and it may become a new immunomodulatory agent in many regions.

In resin transfer molding processes, small clearances exist between the fiber preform and the mold edges, which result in a preferential resin flow in the edge channel and then disrupt the flow patterns during the mold filling stage. A mathematical model including the effect of cavity thickness on resin flow was developed for flow behavior involving the interface between an edge channel and a porous medium. According to the mathematical analysis of momentum equations in a fully developed rectangular duct and formulations of the equivalent edge permeability, comparing with three-dimensional Navier–Stokes equations, the governing equations were modified in the edge channel. The volume of fluid (VOF) method was applied to track the flow front. A simple case is numerically simulated using the modified governing equations. The effects of edge channel width and cavity thickness on flow front and inlet pressure are analyzed, and the evolution characteristics of simulated results are in agreement with the experimental results.

In order to improve the quality of reporting Chinese herbal medicine (CHM) trials, an elaborated checklist was developed according to the basic characteristics of traditional Chinese medicine (TCM) and on the basis of the Reporting Randomized Controlled Trials of Herbal Interventions: An Elaborated CONSORT Statement. This checklist adopted the 22 items of the Herbal CONSORT Statement with some necessary elaboration and modification in the detailed contents of 11 items, among which, the items of Participants, Intervention, Control group and Outcomes were elaborated and modified accordingly to adapt TCM characteristics. Since the design and reporting of trials on Chinese proprietary medicine and herbal decoctions are different in some aspects, the checklist provided two sets of elaborated item 4 (interventions). We hope that authors, journal editors and peer reviewers find this checklist instructive.

p120-catenin (p120), an E-cadherin regulator, has been implicated as central to a series of genetic and epigenetic changes that ultimately lead to tumor progression and metastasis. Ras-related C3 botulinum toxin substrate 1 (Rac1)and p21-activated kinases (PAKs) are effectors of p120. In the present study, we examined the expression of p120, Rac1 and Pak1 using immunohistochemistry in human gastric cancer tissues. Then, we used the gastric cancer SGC7901 and AGS cell lines to explore the possible mechanism of p120, Rac1 and Pak1 in the progress of gastric cancer. Western blotting was used to detect the expression of p120, Rac1 and Pak1 in the two cell lines. Next, p120 was silenced using p120 siRNA or overexpression of p120 by transfection of the plasmid p120 1A into the two cell types, western blotting was used to investigate the expression changes of Rac1 and Pak1. Furthermore, the effects of p120 siRNA-mediated knockdown or overexpression on the proliferation and invasive ability of gastric cancer cells were investigated using wound healing test and Matrigel invasion assays. The results showed that p120 was downregulated in both poorly differentiated group and well differentiated human gastric cancer. However, Rac1 and Pak1 were upregulated in poorly differentiated tissues and remain low in well differentiated gastric cancer tissues. In the two gastric cancer cell lines, although the expression of Rac1 and Pak1 remained unchanged after the p120 knockdown, the expressions of Rac1 and Pak1 protein were decreased after p120 overexpression in both SGC7901 and AGS cells. Furthermore, knockdown of p120 promoted gastric cancer cell proliferation and invasion; overexpression of p120 reduced the proliferation and invasion of gastric cancer cells. In conclusion, based on our results, we speculate that p120 participates in the progress of gastric cancer through regulating Rac1 and Pak1, which provides a potential prevention and a promising therapeutical approach for the patients with gastric cancer.

A set of near infrared high resolution spectral imaging system is set up, the infrared absorption properties of raw cotton and colorless foreign are analyzed through the system, and scheme of polypropylene fiber detection based on the near infrared spectral image is proposed; On this basis, reduce dimensions the spectral images through the principal component analysis, further improve the efficiency of colorless foreign detection. The experimental results show that the spectral images after reducing dimensions can be used to detect colorless or light color raw cotton fiber effectively.

Objective: To investigate respiratory chain complex II deficiency resulted from mutation in succinate dehydrogenase gene (SDH) encoding complex II subunits in China. Methods: An 11-year-old boy was admitted to our hospital. He had a history of progressive psychomotor regression and weakness since the age of 4 years. His cranial magnetic resonance imaging revealed focal, bilaterally symmetrical lesions in the basal ganglia and thalamus, indicating mitochondrial encephalopathy. The activities of mitochondrial respiratory chain enzymes I−V in peripheral leukocytes were determined via spectrophotometry. Mitochondrial DNA and the succinate dehydrogenase A (SDHA) gene were analyzed by direct sequencing. Results: Complex II activity in the leukocytes had decreased to 33.07 nmol/min/mg mitochondrial protein (normal control 71.8 ± 12.9); the activities of complexes I, III, IV and V were normal. The entire sequence of the mitochondrial DNA was normal. The SDHA gene showed two heterozygous frame-shift mutations: c.G117G/del in exon 2 and c.T220T/insT in exon 3, which resulted in stop codons at residues 56 and 81, respectively. Conclusions: We have described the first Chinese case of mitochondrial respiratory chain complex II deficiency, which was diagnosed using enzyme assays and gene analysis. Two novel, compound, frame-shift mutations, c.G117G/del in exon 2 and c.T220T/insT in exon 3 of the SDHA gene, were found in our patient.

Background Argininemia is an autosomal recessive genetic disorder caused by hepatocyte arginase deficiency. It could be detected by blood amino acids analysis (high arginine) and confirmed by molecular diagnosis. The clinical manifestations in patients are similar to cerebral palsy so the diagnosis is usually much delayed. Reports of argininemia from mainland China are few, and genetic analyses have not been reported. Patients and Methods Five Chinese patients with argininemia were investigated. They had progressive spastic tetraplegia, poor physical growth from 1 month to 4 years. When argininemia was found at the ages of 4 to 12 years, four of patients had mental retardation, and three had seizures. Results Elevated blood arginine and significantly decreased erythrocyte arginase activity in five patients confirmed the diagnosis of arginase deficiency. Liver dysfunction was found in four patients, two of whom had mildly elevated blood ammonia levels. Cranial magnetic resonance imaging showed progressive cerebral atrophy in three patients. Six mutations in the ARG1 gene were identified, of which only one (c.703 G>A, p.G235R) in exon 7 has been reported before; c.34 G>T (p.G12X) in exon 1, c.67delG (p.G23fsX31) in exon 2, c.539G>C (p.R180 T) in exon 5, c.374C>T (p.A125 V) in exon 4, and c.646-649del CTCA (p.T215fsX219) in exon 6 were novel mutations. Conclusions Argininemia is one of the few treatable causes of pediatric spastic paraparesis. Early metabolic investigation is very important to reach a diagnosis and better outcome. Five Chinese patients with late-diagnosed argininemia were reported. The mutation spectrum of ARG1 gene should be different from other populations.

Background Argininosuccinate synthetase deficiency (citrullinemia type 1) is a rare autosomal recessive disorder of the urea cycle characterized by elevated concentrations of citrulline, ammonia, and orotic acid, manifesting with acute hyperammonemic crises, usually early in life, with concurrent neurologic deterioration. Only a few cases of citrullinemia type 1 have been documented from mainland China. Prenatal diagnosis has not been performed. Methods A Chinese family affected by citrullinemia type 1 was studied. The proband, a girl, was the second child born to a non-consanguineous couple. Her elder brother died at 19 months due to coma and liver dysfunction of unknown cause. The proband was admitted because of severe mental retardation and lethargy at the age of 15 months. Initial laboratory results revealed hyperammonaemia, hypercitrullinemia (928.771 μmol/L, normal 5.0–25.0 μmol/L) and orotic aciduria, supporting the diagnosis of citrullinemia type 1. Subsequently, the mother presented at 15 weeks of pregnancy seeking for genetic counseling and prenatal diagnosis. ASS1 gene in the blood leukocytes of the family members and amniocytes was performed by direct sequencing. Results On the ASS1 gene of the proband, a novel mutation, T1009C (C337R), and a previously reported mutation G847A (E283K) were identified. Each parent carries one of two mutations. G847A and T1009C mutations were detected in amniocytes, as same as the proband of the family. The result revealed that the fetus was affected by argininosuccinate synthetase deficiency. The parents chose to have a termination of the pregnancy. Conclusions Prenatal diagnosis for citrullinemia type 1 was performed in a Chinese family using gene analysis. T1009C (C337R), a novel mutation of ASS1, was identified.

This study investigated the genetic and enzymological features of Leigh syndrome due to respiratory chain complex deficiency in Chinese patients. The clinical features of 75 patients were recorded. Mitochondrial respiratory chain enzyme activities were determined via spectrophotometry. Mitochondrial gene sequence analysis was performed in 23 patients. Five core pedigrees were investigated via restriction fragment length polymorphism and gene sequencing. Psychomotor retardation (55%), motor regression (20%), weakness (29%), and epilepsy (25%) were the most frequent manifestations. Sixty-four patients (85.3%) had isolated respiratory complex deficiencies: complex I was seen in 28 patients (37.3%); complex II, seven (9.3%); complex III, six (8%); complex IV, ten (13.3%); and complex V, 13 patients (17.3%). Eleven patients (14.7%) had combined complex deficiencies. Mitochondrial DNA mutations were detected in 10 patients. Eight point mutations were found in mitochondrial structural genes: m.4833A > G in ND2, m.10191T > C in ND3, m.12338T > C and m.13513G > A in ND5, m.14502T > C and m.14487T > C in ND6, m.8108A > G in COXII, and m.8993T > G in ATPase6. Three mutations were found in tRNA genes: m.4395A > G in tRNA-Gln, m.10454T > C in tRNA-Arg, and m.5587T > C in tRNA-Ala. One patient and their mother both had the m.12338T > C and m.8993T > C mutations. In conclusion, mitochondrial respiratory chain complex I deficiency and structural gene mutations frequently occur in Chinese Leigh syndrome patients.

Objective Early treatment (growth hormone and nutritional support) improves development in infants with Prader-Willi syndrome. This study aimed to evaluate the nutritional and metabolic condition of nine patients who were diagnosed and treated in early infancy. Methods Nine patients were hospitalized at the age of \xe2\u20ac\xa810 days to 11 months because of severe feeding difficulties, failure to thrive, or developmental delay. The diagnosis of Prader-Willi syndrome was confirmed by fluorescence in situ hybridization or other molecular genetic techniques. Nutritional and metabolic investigations including urinary organic acid analysis, blood amino acid, and acylcarnitine profiles were performed. Results The diagnosis was made at the mean age of 6.3 months. A deletion of the paternal gene in the 15q11-13 region was detected in all patients. Eight patients had ketosis, seven had malnutrition, five had hyperammonemia, three had liver dysfunction, three had low blood cholesterol level, and two had hypoglycemia. All patients had reduction of serum multiple amino acids and free carnitine. Significant arginine deficiency was found in all patients. Six patients had mildly elevated blood long-chain and very long-chain acylcarnitine. After supplementation with l-arginine, medium-chain fatty acids, l-carnitine, and vitamins, all patients responded with improvement of motor development and nutritional conditions. Four patients were almost caught up on physical and psychomotor development. Conclusions Patients with Prader-Willi syndrome are in bad metabolic condition in the early period. Early diagnosis and individual nutritional interventions may improve the nutritional and developmental progress and decrease death rate in infancy.

Summarygp78 is a membrane-anchored ubiquitin ligase mediating the degradation of HMG-CoA reductase (HMGCR) and Insig-1. As a rate-limiting enzyme in cholesterol biosynthesis, HMGCR undergoes rapid sterol-promoted degradation. In contrast, destruction of Insig-1 releases its inhibition on SREBP and stimulates the expression of lipogenic genes. Thus, gp78 has opposite effects on lipid biosynthesis. We here generated liver-specific gp78 knockout (L-gp78−/−) mice and showed that although the degradation of HMGCR was blunted, SREBP was suppressed due to the elevation of Insig-1/-2, and therefore the lipid biosynthesis was decreased. The L-gp78−/− mice were protected from diet-/age-induced obesity and glucose intolerance. The livers of L-gp78−/− mice produced more FGF21, which activated thermogenesis in brown adipocytes and enhanced energy expenditure. Together, the major function of gp78 in liver is regulating lipid biosynthesis through SREBP pathway. Ablation of gp78 decreases the lipid levels and increases FGF21, and is beneficial to patients with metabolic diseases.