SIGLEC10+ macrophages drive gastric cancer progression by suppressing CD8+ T cell function.

Existing immune checkpoint inhibitors focus on activating T cells and show limited effectiveness in gastric cancer (GC). SIGLEC10 is identified as a novel tumor-associated macrophage-related immune checkpoint in other cancer types. However, its immunosuppressive role and clinical significance in GC remain unclear. In this study, we find a dominant expression of SIGLEC10 on CD68+ macrophages in GC. SIGLEC10 can suppress the proliferation and function of tumor-infiltrating CD8+ T cells in vitro via the Akt/P38/Erk signaling pathway. Furthermore, in ex vivo and in vivo models, SIGLEC10 blockade promotes CD8+ T cell effector function. Finally, SIGLEC10+ macrophages are positively correlated with the adverse prognosis of GC. Our study highlights that SIGLEC10 directly suppresses T cell function and serves as a promising target for immunotherapy and suggests SIGLEC10+ macrophages as a novel potential predictor of the clinical prognosis of GC.

CCL19/CCR7 drives regulatory T cell migration and indicates poor prognosis in gastric cancer.

BACKGROUND: Gastric cancer is associated with significant morbidity and mortality in the world. Blocking programmed cell death protein 1 pathway have been approved for the treatment of a variety of tumors and have achieved remarkable clinical therapeutic effects. However, immune checkpoint inhibitors failed to achieve satisfactory results in gastric cancer. There is a need to identify novel immunotherapy targets in gastric cancer. METHODS: We analysed the correlation between Treg cells and CD8 + T cells in gastric cancer samples. We studied the relationship between chemokines and Treg cells or CD8 + T cells in gastric cancer. We compared CCL19/CCR7 expression in gastric cancer patients in TCGA database. We performed transwell experiments to determine the influence of CCL19 on Treg cells and CD8 + T cells migratory capacity. We conducted survival analysis of CCL19 and CCR7 in gastric cancer database. RESULTS: Treg cells show positive correlation with CD8 + T cells in gastric cancer. Treg cell expression was significantly upregulated in tumor tissues. Patients with high FOXP3 expression had worse overall survival than those with low FOXP3 expression. CCL19 had strong correlation with FOXP3 and weak correlation with CD8A. CCL19 had strong impact on the migratory capacity of Treg cells but weak impact on the migratory capacity of CD8 + T cells. Both CCL19 and CCR7 expression were significantly upregulated in gastric cancer tissues. Survival analysis demonstrated that both CCL19 and CCR7 indicate poor prognosis in gastric cancer. CONCLUSIONS: CCL19/CCR7 may be a potential novel therapeutic target in gastric cancer.

Non-small cell lung cancer cells with deficiencies in homologous recombination genes are sensitive to PARP inhibitors.

Lung cancer is the leading cause of cancer death worldwide. PARP inhibitors have become a new line of cancer therapy and a successful demonstration of the synthetic lethality concept. The mechanism and efficacy of PARP inhibitors have been well studied in some cancers, especially homologous recombination (HR)-deficient ovarian cancer and breast cancer, yet such studies are still relatively fewer in lung cancer. Here we found that HR genes are frequently mutated in lung cancer patients, exposing a window for targeted therapies by PARP inhibitors. We depleted BRCA1 and BRCA2 in non-small cell lung cancer (NSCLC) cancer cells and found these cells are hypersensitive to the PARP inhibitor olaparib in cell viability and clonogenic survival assays. Olaparib specifically induces apoptosis in A549 cells with BRCA1 or BRCA2 depletion, as determined by positive Annexin-V staining. In addition, we show that A549 cells with ATM shRNA knockdown are also hypersensitive to Olaparib. In summary, our data support the potential use of PARP inhibitors in NSCLC with HR deficiency.

Distance effect characteristic of the matrix attachment region increases recombinant protein expression in Chinese hamster ovary cells.

OBJECTIVES: Previously, we have found that the matrix attachment region (MAR) may confer a 'distance effect' on transgene expression. This work aims to systematically explore the increased transgene expression in transfected Chinese hamster ovary (CHO) cells due to the characteristics of MAR and its mechanism. RESULTS: Compared with the control vector, 500 and 1000 bp DNA distances between MAR and the cytomegalovirus promoter can increase transgene expression by 1.77- and 1.56-fold, respectively. Meanwhile, transgene expression was not affected when 2000 and 2500 bp spacer DNAs were inserted, but a declining trend was observed when a 1500 bp spacer DNA was inserted. The vector containing a 500 bp DNA distance significantly increased the expression of the enhanced green fluorescent protein, and this increase was not related to transgene copy numbers. CONCLUSIONS: A short DNA distance-containing MAR confers high transgene expression level in transfected CHO cells, but a distance threshold does not exist in the vector system.

Enhancing expression level and stability of transgene mediated by episomal vector via buffering DNA methyltransferase in transfected CHO cells.

Nonviral episomal vectors present attractive alternative vehicles for gene therapy applications. Previously, we have established a new type of nonviral episomal vector-mediated by the characteristic motifs of matrix attachment regions (MARs), which is driven by the cytomegalovirus (CMV) promoter. However, the CMV promoter is intrinsically susceptible to silencing, resulting in declined productivity during long-term culture. In this study, Chinese hamster ovary (CHO) cells and DNA methyltransferase-deficient (Dnmt3a-deficient) CHO cells were transfected with plasmid-mediated by MAR, or CHO cells were treated with the DNA methylation inhibitor 5-Aza-2'-deoxycytidine. Flow cytometry, plasmid rescue experiments, fluorescence in-situ hybridization (FISH), and bisulfite sequencing were performed to observe transgene expression, its state of existence, and the CpG methylation level of the CMV promoter. The results indicated that all DNA methylation inhibitor and methyltransferase deficient cells could increase transgene expression levels and stability in the presence or absence of selection pressure after a 60-generation culture. Plasmid rescue assay and FISH analysis showed that the vector still existed episomally after long-time culture. Moreover, a relatively lower CMV promoter methylation level was observed in Dnmt3a-deficient cell lines and CHO cells treated with 5-Aza-2'-deoxycytidine. In addition, Dnmt3a-deficient cells were superior to the DNA methylation inhibitor treatment regarding the transgene expression and long-term stability. Our study provides the first evidence that lower DNA methyltransferase can enhance expression level and stability of transgenes mediated by episomal vectors in transfected CHO cells.

Somatic mutation of DNAH genes implicated higher chemotherapy response rate in gastric adenocarcinoma patients.

BACKGROUND: The dynein axonemal heavy chain (DNAH) family of genes encode the dynein axonemal heavy chain, which is involved in cell motility. Genomic variations of DNAH family members have been frequently reported in diverse kinds of malignant tumors. In this study, we analyzed the genomic database to evaluate the mutation status of DNAH genes in gastric adenocarcinoma and further identified the significance of mutant DNAH genes as effective molecular biomarkers for predicting chemotherapy response in gastric cancer patients. METHODS: We analyzed the clinical and genomic data of gastric cancer patients published in The Cancer Genome Atlas (TCGA) project. Data on chemotherapy response, overall survival (OS) and chemotherapy-free survival were retrieved. Then, we verified the results via targeted sequencing of gastric cancer patients with similar clinical characteristics but different chemotherapeutic outcomes. RESULTS: In total, 132 gastric adenocarcinoma patients undergoing chemotherapy treatment from TCGA were included in our study. Somatic mutations in all 13 members of the DNAH family of genes were associated with different chemotherapy responses. Compared with patients with wild-type DNAH genes (n = 59), a significantly higher proportion of those with mutations in DNAH genes (n = 73) (55.9% vs 80.8%) responded to chemotherapy (P = 0.002). Moreover, DNAH mutations were correlated with significantly better OS (P = 0.027), chemotherapy-free survival (P = 0.027), fluoropyrimidine-free survival (P = 0.048) and platinum-free survival (P = 0.014). DNAH mutation status was an independent risk factor for OS (P = 0.015), chemotherapy-free survival (P = 0.015) and platinum-free survival (P = 0.011). We identified somatic mutations in 27 (42.2%) of the 64 stage III gastric adenocarcinoma patients receiving fluoropyrimidine-based chemotherapy by targeted exon sequencing with strict screening conditions. In our own cohort, a significantly higher proportion of patients (n = 32) with DNAH mutations than patients with wild-type DNAH genes (n = 32) had a good prognosis (OS > 48 months) (70.4% vs 35.1%) (P = 0.005). CONCLUSIONS: Dynein axonemal heavy chain gene mutations contribute positively to chemotherapy sensitivity in gastric cancer patients.

Regulatory T cells and M2 macrophages present diverse prognostic value in gastric cancer patients with different clinicopathologic characteristics and chemotherapy strategies.

BACKGROUND: Gastric cancer (GC) remains a refractory cancer worldwide. Currently, exploring the differences of the immune status in GC patients with different subgroups might provide promising immunotherapeutic approaches for the treatment of GC. METHODS: In this study, a total of 598 surgically resected FFPE primary gastric cancer samples were assessed for FOXP3, CD163, CD3, CD8, and PD-L1 markers. The correlations between the immune markers expression and clinicopathological features and prognosis were investigated retrospectively. RESULTS: In general, PD-L1, CD3, and CD8 could be regarded as favorable prognostic factors. Our data demonstrated that high infiltration of FOXP3+ Treg indicates better prognosis in stage I-II patients, while the converse outcome was noted in stage III-IV patients. Our data also confirmed different prognostic value in different pathological classifications, chemotherapy strategies, and locations, with or without lymph node metastasis. Also, M2 macrophages indicated poor prognosis in general. However, high M2 macrophage infiltration suggests a favorable prognosis in signet ring cell carcinoma and mucinous adenocarcinoma. Moreover, the prognostic value of the two indices when they are combined is reported. CONCLUSIONS: These results suggested that different immune statuses are exhibited in different subgroups of GC, which may direct further understanding of the immune status of GC as well as provide a further theoretical basis and potential targets for GC immunotherapy.

SV40 intron, a potent strong intron element that effectively increases transgene expression in transfected Chinese hamster ovary cells.

Chinese hamster ovary (CHO) cells have become the most widely utilized mammalian cell line for the production of recombinant proteins. However, the product yield and transgene instability need to be further increased and solved. In this study, we investigated the effect of five different introns on transgene expression in CHO cells. hCMV intron A, adenovirus tripartite leader sequence intron, SV40 intron, Chinese hamster EF-1alpha gene intron 1 and intervening sequence intron were cloned downstream of the eGFP expression cassette in a eukaryotic vector, which was then transfected into CHO cells. qRT-PCR and flow cytometry were used to explore eGFP expression levels. And gene copy number was also detected by qPCR, respectively. Furthermore, the erythropoietin (EPO) protein was used to test the selected more strong intron. The results showed that SV40 intron exhibited the highest transgene expression level among the five compared intron elements under transient and stable transfections. In addition, the SV40 intron element can increase the ratio of positive colonies and decrease the coefficient of variation in transgene expression level. Moreover, the transgene expression level was not related to the gene copy number in stable transfected CHO cells. Also, the SV40 intron induced higher level of EPO expression than IVS intron in transfected CHO cell. In conclusion, SV40 intron is a potent strong intron element that increases transgene expression, which can readily be used to more efficient transgenic protein production in CHO cells.

Identification of a potent MAR element from the human genome and assessment of its activity in stably transfected CHO cells.

Low-level and unstable transgene expression are common issues using the CHO cell expression system. Matrix attachment regions (MARs) enhance transgene expression levels, but additional research is needed to improve their function and to determine their mechanism of action. MAR-6 from CHO chromosomes actively mediates high and consistent gene expression. In this study, we compared the effects of two new MARs and MAR-6 on transgene expression in recombinant CHO cells and found one potent MAR element that can significantly increase transgene expression. Two MARs, including the human CSP-B MAR element and DHFR intron MAR element from CHO cells, were cloned and inserted downstream of the poly(A) site in a eukaryotic vector. The constructs were transfected into CHO cells, and the expression levels and stability of eGFP were detected by flow cytometry. The three MAR sequences can be ranked in terms of overall eGFP expression, in decreasing order, as follows: human CSP-B, DHFR intron MAR element and MAR-6. Additionally, as expected, the three MAR-containing vectors showed higher transfection efficiencies and transient transgene expression in comparison with those of the non-MAR-containing vector. Bioinformatics analysis indicated that the NFAT and VIBP elements within MAR sequences may contribute to the enhancement of eGFP expression. In conclusion, the human CSP-B MAR element can improve transgene expression and its effects may be related to the NFAT and VIBP elements.

Construction strategies for developing expression vectors for recombinant monoclonal antibody production in CHO cells.

Recent years have seen the use of recombinant proteins in the treatment of different diseases. Among them, monoclonal antibodies (mAbs) are currently the fastest growing class of bio-therapeutic recombinant proteins. Chinese hamster ovary (CHO) cells are the most commonly used host cells for production of these recombinant mAbs. Expression vectors determine the expression level and quality of recombinant mAbs. Currently, few construction strategies for recombinant mAbs expression vectors in CHO cells have been developed, including monocistronic vector, multiple-promoter expression vector, and tricistronic vector mediated by internal ribosome entry site (IRES) or Furin-2A element. Among them, Furin-2A-mediated vector is an effective approach due to advantages of high "self-cleavage" efficiency, and equal expression of light and heavy chains from a single open reading frame. Here, we have reviewed the progress in development of different strategies for constructing recombinant mAb expression vectors in CHO cells and its potential advantages and disadvantages.

[Effects of Different Promoters and MAR Combinations on Transgene Expression of Recombinant CHO Cells].

OBJECTIVE: To analyze the effects of different promoters and matrix attachment region (MAR) on the expression of transgene in Chinese hamster ovary (CHO) cells. METHODS: The expression vector was constructed by the combination of beta globin MAR (gMAR) with the human cytomegalovirus immediate-early promoter (CMV-IE) and simian virus 40 (SV40) promoter. These vectors were transfected into CHO cells,after 48 h,the transient expression of enhanced green fluorescent protein (eGFP) was observed; G418 was used to screen stably transformed cell lines,and the expression level of eGFP in CHO cells was analyzed by flow cytometry. The relative copy numbers of eGFP were analyzed by qPCR. RESULTS: Without gMAR expression vector,the expression of eGFP which was driven by CMV-IE promoter was stronger than that of SV40 promoter; gMAR could increase the expression level of eGFP driven by CMV-IE promoter,but did not show any enhancement in SV40 promoter. The expression level of eGFP which containing gMAR on both sides was stronger than that of gMAR on one side driven by CMV-IE promoter; After G418 screening,the expression level of eGFP containing gMAR driven by SV40 promoter wasunstable,the fluorescence gradually weakened,therefore,we only analyzed the expression vector stably expressing the eGFP gene driven by CMV-IE promoter by flow cytometry and qPCR. Compared with the expression vector without gMAR containing CMV-IE promoter,flow cytometry showed that the expression levels of eGFP on one and both sides with gMAR were increased by 9.85-fold and 12.94-fold,respectivley; The result of qPCR showed that the copy number of the eGFP gene without gMAR was set to 1,the copy number of the eGFP gene in the expression vector driven by CMV-IE with gMAR on one side and both sides were 3.68-fold and 9.25-fold,respectively. CONCLUSION: The activity of CMV-IE promoter is stronger than that of SV40 promoter. gMAR can enhance the expression levels of transgene,which may be related to the increase of gene copy number.

[Medication regularity and potential targets of Professor XU Jing-fan's prescription for treating ulcerative colitis].

Ulcerative colitis (UC) is a chronic nonspecific inflammation mainly involving rectum and colon mucosa, which seriously affects the health and quality of life of patients, and is listed as one of modern refractory diseases by WHO. Professor XU Jing-fan, a great master of traditional Chinese medicine, has accumulated rich experiences in the treatment of UC. The study collected Professor XU's 77 prescriptions of treating UC, analyzed the frequency of traditional Chinese medicines and there categories, and investigated the medication regularity by the system clustering method. The findings showed that the most frequently used drugs were clearing-heat herbs, which were followed by hemostatic herbs, excreting-dampness herbs, improving-digestion herbs and tonifying-Qi herbs. At the same time, the commonly combined drugs were excavated. Finally, in order to analyze potential molecular targets of the frequently used herbs, GO enrichment analysis and KEGG signal pathway enrichment analysis were performed with bioinformatics analysis tool for molecular mechanism of traditional Chinese medicine (BATMAN-TCM). The results indicated that Chinese herbal compounds may treat UC by activating PPAR-γ pathway and regulating intestinal inflammation. The exact mechanisms shall be verified through subsequent molecular biological experiments.

The EF-1α promoter maintains high-level transgene expression from episomal vectors in transfected CHO-K1 cells.

In our previous study, we demonstrated that episomal vectors based on the characteristic sequence of matrix attachment regions (MARs) and containing the cytomegalovirus (CMV) promoter allow transgenes to be maintained episomally in Chinese hamster ovary (CHO) cells. However, the transgene expression was unstable and the number of copies was low. In this study, we focused on enhancers, various promoters and promoter variants that could improve the transgene expression stability, expression magnitude (level) and the copy number of a MAR-based episomal vector in CHO-K1 cells. In comparison with the CMV promoter, the eukaryotic translation elongation factor 1 α (EF-1α, gene symbol EEF1A1) promoter increased the transfection efficiency, the transgene expression, the proportion of expression-positive clones and the copy number of the episomal vector in long-term culture. By contrast, no significant positive effects were observed with an enhancer, CMV promoter variants or CAG promoter in the episomal vector in long-term culture. Moreover, the high-expression clones harbouring the EF-1α promoter tended to be more stable in long-term culture, even in the absence of selection pressure. According to these findings, we concluded that the EF-1α promoter is a potent regulatory sequence for episomal vectors because it maintains high transgene expression, transgene stability and copy number. These results provide valuable information on improvement of transgene stability and the copy number of episomal vectors.

Clinical pharmacist interventions on a Chinese neurology ward: a 2-year service evaluation.

A prospective observational study of clinical pharmacist interventions was conducted over a 2-year period from November 2012 to October 2014 to evaluate the clinical activity of pharmacists in the care they provide to patients and to promote safe and effective medication therapy by quantifying medicine-related interventions on a Chinese neurology ward. All pharmacist interventions made in the department of neurology were recorded, categorized, and assessed for potential patient harm if the intervention had not taken place. The quantity, outcomes, and potential severity of clinical pharmacists' interventions were recorded. 619 interventions were made in 385 patients over the 2-year observational period. The mean severity of potential harm assessment was 3.7 (1.12), range 0.8 - 7.0. 87 of the 619 interventions (14.0%) were classified as medication errors. The results of the clinical pharmacist intervention study demonstrated that pharmacists play an important role in the care of neurological patients by improving patient care and reducing clinical risk.

[Simultaneous determination of phenolic acids in Guanxinning injection by HPLC].

This paper was aim to determine five phenolic acids, sodium danshensu (SD), protocatechuic aldehyde (PA), rosmarinic acid (RA), lithospermic acid (LA) and salvianolic acid B (SAB), in Guanxinning injection. In the test, Kromasil C18 column (4.6 mm x 250 mm, 5 µm) was adopted, with acetonitrile-3% formic acid solution as the mobile phase for gradient elution. The flow rate was 1 mL · min, the column temperature was 30 °C and the detection wavelength was 280 nm. According to the results of the test, SD, PA, RA, LA and SAB showed good linear relations between peak areas and sample sizes in 0.006 06-4.04 (r = 0.999 3), 0.006 15-4.10 (r = 0.999 4), 0.005 94-3.96 (r = 0.999 3), 0.006 06-4.04(r = 0.999 1) and 0.006 09-4.06 (r = 0.999 2) µg, respectively. The average recoveries (n = 6) were 98.9% (RSD 0.75%), 98.1% (RSD 1.2%), 100% (RSD 0.77%), 98.7% (RSD 1.7%), 102% (RSD 0.68%), respectively. The above 5 components were determined in 13 batches of samples by using the established method. The method was simple, accurate and highly reproducible that it could be used for quality control of the components in Guanxinning injection.

[Discussion about risk and management of Chinese patent medicine with double identity].

Chinese patent medicine with double identity was a special phenomenon, and many preparations not only were prescription drugs but also over the counter ( OTC) drugs, which brought a lot of trouble. Based on statistics of list of OTC medicines of CFDA, related varieties, route of administration and functions of these drugs were searched. The causes of insufficient were analyzed and the potential risk was investigated. To ensure the safety of drug usage for the patient, risk management system should be set up by improving the technical requirements for registration, improving the drug labels and manuals, playing the role of pharmacists in pharmacy services and raising awareness of doctor and patient for these drugs.

CD177 drives the transendothelial migration of Treg cells enriched in human colorectal cancer.

Objectives: Regulatory T (Treg) cells regulate immunity in autoimmune diseases and cancers. However, immunotherapies that target tumor-infiltrating Treg cells often induce unwanted immune responses and tissue inflammation. Our research focussed on exploring the expression pattern of CD177 in tumor-infiltrating Treg cells with the aim of identifying a potential target that can enhance immunotherapy effectiveness. Methods: Single-cell RNA sequencing (scRNA-seq) data and survival data were obtained from public databases. Twenty-one colorectal cancer patient samples, including fresh tumor tissues, peritumoral tissues and peripheral blood mononuclear cells (PBMCs), were analysed using flow cytometry. The transendothelial activity of CD177+ Treg cells was substantiated using in vitro experiments. Results: ScRNA-seq and flow cytometry results indicated that CD177 was exclusively expressed in intratumoral Treg cells. CD177+ Treg cells exhibited greater activation status and expressed elevated Treg cell canonical markers and immune checkpoint molecules than CD177- Treg cells. We further discovered that both intratumoral CD177+ Treg cells and CD177-overexpressing induced Treg (iTreg) cells had lower levels of PD-1 than their CD177- counterparts. Moreover, CD177 overexpression significantly enhanced the transendothelial migration of Treg cells in vitro. Conclusions: These results demonstrated that Treg cells with higher CD177 levels exhibited an enhanced activation status and transendothelial migration capacity. Our findings suggest that CD177 may serve as an immunotherapeutic target and that overexpression of CD177 may improve the efficacy of chimeric antigen receptor T (CAR-T) cell therapy.

Nuclear Tubulin Enhances CXCR4 Transcription and Promotes Chemotaxis Through TCF12 Transcription Factor in human Hematopoietic Stem Cells.

Tubulins are cytoskeleton components in all eukaryotic cells and play crucial roles in various cellular activities by polymerizing into dynamic microtubules. A subpopulation of tubulin has been shown to localize in the nucleus, however, the function of nuclear tubulin remains largely unexplored. Here we report that microtubule depolymerization specifically upregulates surface CXCR4 expression in human hematopoietic stem cells (HSCs). Mechanistically, microtubule depolymerization results in accumulation of tubulin subunits in the nucleus, leading to elevated CXCR4 transcription and increased chemotaxis of human HSCs. Treatment with microtubule stabilizer Epothilone B strongly suppresses the phenotypes induced by microtubule depolymerizing agents in human HSCs. Furthermore, chromatin immunoprecipitation assay reveals an increased binding of nuclear tubulin and TCF12 transcription factor at the CXCR4 promoter region. Depletion of TCF12 significantly suppresses microtubule depolymerization mediated upregulation of CXCR4 surface expression. These results demonstrate a previously unknown function of nuclear tubulin in regulating gene transcription through TCF12. New strategy targeting nuclear tubulin-TCF12-CXCR4 axis may be applicable to enhance HSC transplantation.

Qingre Xingyu recipe exerts inhibiting effects on ulcerative colitis development by inhibiting TNFα/NLRP3/Caspase-1/IL-1ß pathway and macrophage M1 polarization.

As a chronic inflammatory bowel disease, ulcerative colitis (UC) imposes a significant burden on public healthcare worldwide due to its increasing morbidity. Chinese medicines are regarded as potent therapeutic agents for UC treatment with minimal side effects. In the present study, we sought to determine the novel role of a traditional medicine Qingre Xingyu (QRXY) recipe in the development of UC and aimed to contribute to the currently available knowledge about UC by exploring the downstream mechanism of QRXY recipe in UC. Mouse models of UC were established by injections with dextran sulphate sodium (DSS), where the expression of tumor necrosis factor-alpha (TNFα), NLR family pyrin domain containing 3 (NLRP3), and interleukin-1ß (IL-1ß) was determined followed by an analysis of their interactions. The DSS-treated NLRP3 knockout (-/-) Caco-2 cell model was successfully constructed. The in vitro and in vivo effects of the QRXY recipe on UC were investigated with the determination of disease activity index (DAI), histopathological scores, transepithelial electrical resistance, FITC-dextran, as well as cell proliferation and apoptosis. In vivo and in vitro experiments indicated that the QRXY recipe reduced the degree of intestinal mucosal injury of UC mice and functional damage of DSS-induced Caco-2 cells by inhibition of the TNFα/NLRP3/caspase-1/IL-1ß pathway and M1 polarization of macrophages, and TNFα overexpression or NLRP3 knockdown could counterweigh the therapeutic effects of QRXY recipe. To conclude, our study elicited that QRXY inhibited the expression of TNFα and inactivated the NLRP3/Caspase-1/IL-1ß pathway, thereby alleviating intestinal mucosal injury and relieving UC in mice.

[Effect of 18alpha-glycyrrhizic acid and 18beta-glycyrrhizic acid on P-gp function and expression in Caco-2 cells].

OBJECTIVE: The aim of the present study was to evaluate the modulating effect of glycyrrhizic acid C-18 epimers, 18alpha-glycyrrhizic acid (alpha-GL) and 18beta-glycyrrhizic acid (beta-GL) on both P-glycoprotein (P-gp) activity and expression in Caco-2 cell. METHOD: The effects of P-gp activity were analyzed by rhodamine (Rhd 123) accumulation test, and those of P-gp expression were analyzed by flow cytometry and real-time PCR. RESULT: At middle and high concentrations (10, 60 micromol x L(-1)), alpha-GL inhibited the function of P-gp and with on dose dependent while beta-GL induced the function of P-gp at three test concentrations with no dose dependent too. At middle and high concentrations (10, 60 micromol x L(-1)), alpha-GL down-regulated the expression of MDR1 mRNA. At high concentrations (60 micromol x L(-1)), beta-GL up-regulated the expression of MDR1 mRNA; At high concentrations (60 micromol x L(-1)), beta-GL induced the expression of P-gp protein while alpha-GL has no effect on the expression of P-gp protein at three test concentrations. CONCLUSION: The effects of alpha-GL and beta-GL on the expression of MDR1 mRNA and CYP3A mRNA showed the same trend. The character that epimers of GL act on CYP3A and P-gp show similar stereo selectivity whether relate to PXR need further study.