[Expression and Prognostic Impact of HK2 in Bone Marrow of Patients with Newly Diagnosed Acute Myeloid Leukemia].

OBJECTIVE: To detect the expression level of HK2 gene in the bone marrow of newly diagnosed patients with acute myeloid leukemia (AML) and investigate its influence on the clinical characteristics and prognosis. METHODS: The expression level of HK2 gene in the bone marrow of 90 newly diagnosed patients with AML that accompanying clinical characteristics and survival status were detected by RT-qPCR, and compared with 18 allogeneic hematopoietic stem cell transplantation (allo-HSCT) donors. The Chi-square test, Kaplan-Meier survival analysis, and Cox proportional hazards regression model were used to analyze the correlation of HK2 expression level with clinical characteristics and prognosis. RESULTS: Compared with allo-HSCT donors, the HK2 expression was significantly increased in newly diagnosed AML patients (P <0.01). Compared with patients with total response (OR, complete response + complete response with incomplete hematologic recovery) after 2 courses of induction chemotherapy, the expression of HK2 in patients without OR was significantly increased (P <0.05). There was a significant difference in the relative expression of HK2 between patients with and without OR after 2 courses of induction therapy (P <0.001). The median survival time of patients with high expression of HK2 was significantly shorter than that of patients with low expression of HK2 (P <0.05). The multivariate Cox proportional hazards regression analysis showed that prognostic stratification, the expression level of HK2, and whether two courses of induction therapy achieved OR were independent factors affecting the prognosis of AML patients (P <0.05). CONCLUSIONS: Compared with allo-HSCT donors, the expression level of HK2 gene is increased in the bone marrow of newly diagnosed AML patients. The prognosis of patients with high expression of HK2 is poor. The expression level of HK2 is an independent factor affecting the prognosis of AML patients.

Electroacupuncture alleviates postoperative pain through inhibiting neuroinflammation via stimulator of interferon genes/type-1 interferon pathway.

OBJECTIVE: This work explores the impact of electroacupuncture (EA) on acute postoperative pain (APP) and the role of stimulator of interferon genes/type-1 interferon (STING/IFN-1) signaling pathway modulation in the analgesic effect of EA in APP rats. METHODS: The APP rat model was initiated through abdominal surgery and the animals received two 30 min sessions of EA at bilateral ST36 (Zusanli) and SP6 (Sanyinjiao) acupoints. Mechanical, thermal and cold sensitivity tests were performed to measure the pain threshold, and electroencephalograms were recorded in the primary somatosensory cortex to identify the effects of EA treatment on APP. Western blotting and immunofluorescence were used to examine the expression and distribution of proteins in the STING/IFN-1 pathway as well as neuroinflammation. A STING inhibitor (C-176) was administered intrathecally to verify its role in EA. RESULTS: APP rats displayed mechanical and thermal hypersensitivities compared to the control group (P < 0.05). APP significantly reduced the amplitude of θ, α and γ oscillations compared to their baseline values (P < 0.05). Interestingly, expression levels of proteins in the STING/IFN-1 pathway were downregulated after inducing APP (P < 0.05). Further, APP increased pro-inflammatory factors, including interleukin-6, tumor necrosis factor-α and inducible nitric oxide synthase, and downregulated anti-inflammatory factors, including interleukin-10 and arginase-1 (P < 0.05). EA effectively attenuated APP-induced painful hypersensitivities (P < 0.05) and restored the θ, α and γ power in APP rats (P < 0.05). Meanwhile, EA distinctly activated the STING/IFN-1 pathway and mitigated the neuroinflammatory response (P < 0.05). Furthermore, STING/IFN-1 was predominantly expressed in isolectin-B4- or calcitonin-gene-related-peptide-labeled dorsal root ganglion neurons and superficial laminae of the spinal dorsal horn. Inhibition of the STING/IFN-1 pathway by intrathecal injection of C-176 weakened the analgesic and anti-inflammatory effects of EA on APP (P < 0.05). CONCLUSION: EA can generate robust analgesic and anti-inflammatory effects on APP, and these effects may be linked to activating the STING/IFN-1 pathway, suggesting that STING/IFN-1 may be a target for relieving APP. Please cite this article as: Ding YY, Xu F, Wang YF, Han LL, Huang SQ, Zhao S, Ma LL, Zhang TH, Zhao WJ, Chen XD. Electroacupuncture alleviates postoperative pain through inhibiting neuroinflammation via stimulator of interferon genes/type-1 interferon pathway. J Integr Med. 2023; 21(5): 496-508.

Preventive effect of LCZ696 on hypoxic pulmonary hypertension in rats via regulating the PI3K/AKT signaling pathway.

Hypoxic pulmonary hypertension (HPH) is a devastating disease worldwide; however, effective therapeutic drugs are lacking. This study investigated the effects and underlying mechanisms of LCZ696 treatment on hypoxia-induced pulmonary hypertension. Male Sprague-Dawley (SD) rats were kept in a hypobaric chamber with an oxygen concentration of 5% for 4 weeks. Rats were treated with either LCZ696 (18 mg/kg, 36 mg/kg, and 72 mg/kg) or sildenafil. The mean pulmonary artery pressure (mPAP), right ventricle hypertrophy index (RVHI), and lung system index were measured. Hematoxylin-eosin (HE) staining, Masson staining, and immunofluorescence staining were used for histological analysis. Enzyme linked immunosorbent assay (ELISA) kits were used to determine the concentrations of inflammatory and hypoxia-related factors. Western blotting was used to examine the expression of apoptotic and PI3K/AKT signaling pathway proteins in rat lung tissue. Hypoxia increased mPAP, RVHI, and lung system index and induced pulmonary vascular remodeling, pulmonary arteriomyosis, and pulmonary artery fibrosis. LCZ696 treatment reduced the increase in mPAP, RVHI, and the lung system index and ameliorated the induced pathological changes. Hypoxia upregulated expression of NF-kB, TNF-α, IL-6, HIF-1α, and Vascular endothelial growth factor (VEGF), decreased the ratio of Bax/Bcl-2, and activated the PI3K/AKT signaling pathway in lung tissue, and these effects were partially reversed by treatment with LCZ696. These results demonstrated that LCZ696 can ameliorate hypoxia-induced HPH by suppressing apoptosis, inhibiting the inflammatory response, and inhibiting the PI3K/AKT signaling pathway. It provides a reference for clinical rational drug use and lays a foundation for the study of HPH therapeutic drugs.

Whole-Genome Sequencing of 5-Hydroxymethylcytosine at Base Resolution by Bisulfite-Free Single-Step Deamination with Engineered Cytosine Deaminase.

The epigenetic modification 5-hydroxymethylcytosine (5hmC) plays a crucial role in the regulation of gene expression. Although some methods have been developed to detect 5hmC, direct genome-wide mapping of 5hmC at base resolution is still highly desirable. Herein, we proposed a single-step deamination sequencing (SSD-seq) method, designed to precisely map 5hmC across the genome at single-base resolution. SSD-seq takes advantage of a screened engineered human apolipoprotein B mRNA-editing catalytic polypeptide-like 3A (A3A) protein, known as eA3A-v10, to selectively deaminate cytosine (C) and 5-methylcytosine (5mC) but not 5hmC. During sequencing, the deaminated C and 5mC are converted to uracil (U) and thymine (T), read as T in the sequencing data. However, 5hmC remains unaffected by eA3A-v10 and is read as C during sequencing. Consequently, the presence of C in the sequence reads indicates the original 5hmC. We applied SSD-seq to generate a base-resolution map of 5hmC in human lung tissue. Our findings revealed that 5hmC was predominantly localized to CpG dinucleotides. Furthermore, the base-resolution map of 5hmC generated by SSD-seq demonstrated a strong correlation with prior ACE-seq results. The advantages of SSD-seq are its single-step process, absence of bisulfite treatment or DNA glycosylation, cost effectiveness, and ability to detect and quantify 5hmC directly at single-base resolution.

Structural Characterization and Assessment of Anti-Inflammatory Activities of Polyphenols and Depsidone Derivatives from Melastoma malabathricum subsp. normale.

The roots of Melastoma malabathricum subsp. normale (D. Don) Karst. Mey have been used in traditional ethnic medicine systems in China to treat inflammation-triggered ailments, such as trauma, toothache, and fever. Therefore, the aim of this study is to screen for compounds with anti-inflammatory activity in the title plant. The extract of M. malabathricum subsp. normale roots was separated using various chromatographic methods, such as silica gel, ODS C18, MCI gel, and Sephadex LH-20 column chromatography, as well as semi-preparative HPLC. One new complex tannin, named whiskey tannin D (1), and an undescribed tetracyclic depsidone derivative, named guanxidone B (2), along with nine known polyphenols (2-10) and three known depsidone derivatives (12-14) were obtained from this plant. The structures of all compounds were elucidated by extensive NMR and CD experiments in conjunction with HR-ESI-MS data. All these compounds were isolated from this plant for the first time. Moreover, compounds 1-4, 8, and 10-14 were obtained for the first time from the genus Melastoma, and compounds 1, 2, and 11-14 have not been reported from the family Melastomataceae. This is the first report of complex tannin and depsidone derivatives from M. malabathricum subsp. normale, indicating their chemotaxonomic significance to this plant. Compounds 1-12 were investigated for their anti-inflammatory activities on the production of the nitric oxide (NO) in lipopolysaccharide (LPS)-stimulated RAW264.7 cells, and compounds 1, 11, and 12 showed anti-inflammatory activities with IC50 values of 6.46 ± 0.23 µM, 8.02 ± 0.35 µM, and 9.82 ± 0.43 µM, respectively. The structure-activity relationship showed that the catechin at glucose C-1 in ellagitannin was the key to its anti-inflammatory activity, while CH3O- at C-16 of aromatic ring A in depsidone derivatives had little effect on its anti-inflammatory activity. The study of structure-activity relationships is helpful to quickly discover new anti-inflammatory drugs. The successful isolation and structure identification of these compounds, especially complex tannin 1, not only provide materials for the screening of anti-inflammatory compounds, but also provide a basis for the study of chemical taxonomy of the genus Melastoma.

Structural Characterization and Assessment of Anti-Inflammatory and Anti-Tyrosinase Activities of Polyphenols from Melastoma normale.

Polyphenols, widely distributed in the genus Melastoma plants, possess extensive cellular protective effects such as anti-inflammatory, anti-tyrosinase, and anti-obesity, which makes it a potential anti-inflammatory drug or enzyme inhibitor. Therefore, the aim of this study is to screen for the anti-inflammatory and enzyme inhibitory activities of compounds from title plant. Using silica gel, MCI, ODS C18, and Sephadex LH-20 column chromatography, as well as semipreparative HPLC, the extract of Melastoma normale roots was separated. Four new ellagitannins, Whiskey tannin C (1), 1-O-(4-methoxygalloyl)-6-O-galloyl-2,3-O-(S)-hexahydroxydiphenoyl-ß-d-glucose (2), 1-O-galloyl-6-O-(3-methoxygalloyl)-2,3-O-(S)-hexahydroxydiphenoyl-ß-d-glucose (3), and 1-O-galloyl-6-O-vanilloyl-2,3-O-(S)-hexahydroxydiphenoyl-ß-d-glucose (4), along with eight known polyphenols were firstly obtained from this plant. The structures of all isolates were elucidated by HRMS, NMR, and CD analyses. Using lipopolysaccharide (LPS)-stimulated RAW2 64.7 cells, we investigated the anti-inflammatory activities of compounds 1-4, unfortunately, none of them exhibit inhibit nitric oxide (NO) production, their IC50 values are all > 50 µM. Anti-tyrosinase activity assays was done by tyrosinase inhibition activity screening model. Compound 1 showed weak tyrosinase inhibitory activity with IC50 values of 426.02 ± 11.31 µM. Compounds 2-4 displayed moderate tyrosinase inhibitory activities with IC50 values in the range of 124.74 ± 3.12-241.41 ± 6.23 µM. The structure-activity relationships indicate that hydroxylation at C-3', C-4', and C-3 in the flavones were key to their anti-tyrosinase activities. The successful isolation and structure identification of ellagitannin provide materials for the screening of anti-inflammatory drugs and enzyme inhibitors, and also contribute to the development and utilization of M. normale.

Ellagitannins and Oligomeric Proanthocyanidins of Three Polygonaceous Plants.

The aim of this study was to characterize hydrolyzable tannins in Polygonaceous plants, as only a few plants have previously been reported to contain ellagitannins. From Persicaria chinensis, a new hydrolyzable tannin called persicarianin was isolated and characterized to be 3-O-galloyl-4,6-(S)-dehydrohexahydroxydiphenoyl-d-glucose. Interestingly, acid hydrolysis of this compound afforded ellagic acid, despite the absence of a hexahydroxydiphenoyl group. From the rhizome of Polygonum runcinatum var. sinense, a large amount of granatin A, along with minor ellagitannins, helioscpoinin A, davicratinic acids B and C, and a new ellagitannin called polygonanin A, were isolated. Based on 2D nuclear magnetic resonance (NMR) spectroscopic examination, the structure of polygonanin A was determined to be 1,6-(S)-hexahydroxydiphenoyl-2,4-hydroxychebuloyl-ß-d-glucopyranose. These are the second and third hydrolyzable tannins isolated from Polygonaceous plants. In addition, oligomeric proanthocyanidins of Persicaria capitatum and P. chinensis were characterized by thiol degradation. These results suggested that some Polygonaceous plants are the source of hydrolyzable tannins not only proanthocyanidins.

Association Between Initiation, Intensity, and Cessation of Smoking and Mortality Risk in Patients With Cardiovascular Disease: A Cohort Study.

Objectives: To examine the effect of smoking status, smoking intensity, duration of smoking cessation and age of smoking initiation on the risk of all-cause and cause-specific mortality among cardiovascular disease (CVD) patients. Design: A population-based prospective cohort study. Setting: The National Health Interview Survey (NHIS) in the U.S. that were linked to the National Death Index (NDI). Participants: 66,190 CVD participants ≥ 18 years of age who were interviewed between 1997 and 2013 in the NHIS linked to the NDI through December 31, 2015. Outcome Measures: The primary outcome was all-cause mortality and the secondary outcome was cause-specific mortality including CVD mortality and cancer mortality. Results: During the mean follow-up of 8.1 years, we documented 22,518 deaths (including 6,473 CVD deaths and 4,050 cancer deaths). In the overall CVD population, former and current smokers had higher risk of all-cause (Former smokers: hazard ratios (HRs), 1.26; 95% confidence interval (CI), 1.21-1.31, P < 0.001; Current smokers: HRs, 1.96; 95%CI, 1.86-2.07, P < 0.001), CVD (Former smokers: HRs, 1.12; 95%CI, 1.05-1.21, P = 0.001; Current smokers: HRs, 1.80; 95%CI, 1.64-1.97, P < 0.001) and cancer mortality (Former smokers: HRs, 1.49; 95%CI, 1.35-1.64, P < 0.001; Current smokers: HRs, 2.78; 95%CI, 2.49-3.09, P < 0.001) than never smokers. Furthermore, similar results were observed when the study subjects were stratified according to the type of CVD. Among current smokers, the risk for cancer mortality increased as the daily number of cigarettes increased, regardless of the specific type of CVD. However, the association of the risk for all-cause and CVD mortality with smoking intensity did not present a dose-response relationship. In participants with angina pectoris or stroke, smoking intensity was inversely associated with deaths from CVD. In addition, the risk for all-cause, CVD and cancer mortality declined as years of smoking cessation increased. Finally, the relative risk of all-cause mortality was not significantly different in individuals with a younger age of smoking initiation. Conclusions: CVD patients who are smokers have an increased risk of all-cause, CVD and cancer mortality, and the risk decreases significantly after quitting smoking. These data further provide strong evidence that supports the recommendation to quit smoking for the prevention of premature deaths among individuals with CVD.

The Akt/glycogen synthase kinase-3ß pathway participates in the neuroprotective effect of interleukin-4 against cerebral ischemia/reperfusion injury.

Interleukin-4 (IL-4) has a protective effect against cerebral ischemia/reperfusion injury. Animal experiments have shown that IL-4 improves the short- and long-term prognosis of neurological function. The Akt (also called protein kinase B, PKB)/glycogen synthase kinase-3ß (Akt/GSK-3ß) signaling pathway is involved in oxidative stress, the inflammatory response, apoptosis, and autophagy. However, it is not yet clear whether the Akt/GSK-3ß pathway participates in the neuroprotective effect of IL-4 against cerebral ischemia/reperfusion injury. In the present study, we established a cerebral ischemia/reperfusion mouse model by middle cerebral artery occlusion for 60 minutes followed by a 24-hour reperfusion. An IL-4/anti-IL-4 complex (10 µg) was intraperitoneally administered 30 minutes before surgery. We found that administration of IL-4 significantly alleviated the neurological deficits, oxidative stress, cell apoptosis, and autophagy and reduced infarct volume of the mice with cerebral ischemia/reperfusion injury 24 hours after reperfusion. Simultaneously, IL-4 activated Akt/GSK-3ß signaling pathway. However, an Akt inhibitor LY294002, which was injected at 15 nmol/kg via the tail vein, attenuated the protective effects of IL-4. These findings indicate that IL-4 has a protective effect on cerebral ischemia/reperfusion injury by mitigating oxidative stress, reducing apoptosis, and inhibiting excessive autophagy, and that this mechanism may be related to activation of the Akt/GSK-3ß pathway. This animal study was approved by the Animal Ethics Committee of Renmin Hospital of Wuhan University, China (approval No. WDRY2017-K037) on March 9, 2017.

Risk of non-melanoma skin cancer for rheumatoid arthritis patients receiving TNF antagonist: a systematic review and meta-analysis.

OBJECTIVE: Tumor necrosis factor inhibitors (anti-TNF) have become the standard treatment for rheumatoid arthritis (RA). However, evidence is inconsistent as to whether RA patients with anti-TNF are associated with an increased risk of non-melanoma skin cancer (NMSC) compared with those without anti-TNF. We performed a systematic review and meta-analysis to evaluate the risk of NMSC in patients with anti-TNF drugs compared with those without anti-TNF. METHODS: We did a systematic literature search with PubMed, EMBASE, and the Cochrane Library from inception to April 1, 2019. Prospective observational studies were eligible for inclusion if they included any of the approved anti-TNF drugs and reported the risk estimates and 95% confidence interval (95% CI) of NMSC associated with anti-TNF in RA patients. Pooled relative risks (RRs) and 95% CIs were calculated using a fixed-effects model. To assess the heterogeneity and risk of publication bias, we respectively conducted the subgroup and sensitivity analysis, funnel plot, Begg's and Egger's test. RESULTS: The present meta-analysis included six studies with 123,031 patients. Compared with RA patients without anti-TNF, patients with anti-TNF drugs were associated with an increased risk of NMSC (RR 1.28, 95% CI 1.19 to 1.38; I2 = 45.6%, P = 0.056), especially squamous cell skin cancer (SCC) (RR 1.30, 95% CI 1.09 to 1.54; I2 = 0%, P = 0.854), but not basal cell skin cancer (RR 1.13, 95% CI 0.97 to 1.31; I2 = 0%, P = 0.555). Sensitivity and subgroup analysis confirmed the robustness of the primacy results. There was no evidence of publication bias with Begg's and Egger's test or by inspection of the funnel plot. CONCLUSIONS: These results suggest that RA patients treated with anti-TNF are at an increased risk of NMSC, especially SCC. However, this association in RA urgently needs the more clinical studies and basic researches to further validate.Key Points• Rheumatoid arthritis patients treated with tumor necrosis factor inhibitors are associated with a higher risk of non-melanoma skin cancer compared to those patients treated without tumor necrosis factor inhibitors. Hence, tumor necrosis factor inhibitors may be avoided in rheumatoid arthritis patients who are at high risk of non-melanoma skin cancer.• Of note, rheumatoid arthritis patients who were treated for tumor necrosis factor inhibitors compared with patients who were not treated for tumor necrosis factor inhibitors were at significantly increased risk of squamous cell skin cancer, but were not at increased risk of basal cell skin cancer. Therefore, use of tumor necrosis factor inhibitors in rheumatoid arthritis patients should be paid attention to the occurrence of squamous cell skin cancer.

[Effects of MiR-221-Mediated Wnt/ß- Catenin Signaling Pathway on Biological Activity of Childhood Acute Lymphoblastic Leukemia Cells].

OBJECTIVE: To study the effects of miR-221 on the biological activity of childhood acute lymphoblastic leukemia cells and its mechanism. METHODS: Bone marrow mononuclear cells (BMNC) were isolated from bone marrow samples of ALL children diagnosed in our hospital from May 2018 to November 2018. The cells were divided into control group, miR-221-NC group and miR-221 group. After transfection according to the instructions of Lipofectamine 2000 kit, the levels of miR-221 in each group were detected by RT-PCR. Flow cytometry was used to detect the effects of miR-221 on cell cycle and apoptosis. Transwell assay was used to detect cell migration and invasion. Western blot was used to detect the effects of miR-221 on proliferating cell nuclear antigen (PCNA), Caspase 3, Cyclin D1 and MMP-9 proteins in BMNC. Luciferase reporter gene assay was used to detect the targeting relationship between miR-221 and Wnt gene. RESULT: The expression level of miR-221 in the miR-221 group was significantly higher than that in the control group and the miR-221-NC group (P＜0.05). MTT assay showed that, after transfection for 2, 3, 4 and 5 days, the cell proliferation level in miR-221 group was significantly lower than that in the control group and the miR-221-NC group (P＜0.05). The cell ratio of G0/G1 phase was (73.25±8.1)% in the miR-221 group, which was significantly higher than that in the control group and the miR-221-NC group (P＜0.05); moreover, the cell ratio of S phase in the miR-221 group was (12.37±1.6)%，which was significantly lower than that in the control group and the miR-221-NC group (P＜0.05). The percentage of apoptotic cells in the miR-221 group was (24.68±3.87)%, which was significantly higher than that in the control group and the miR-221-NC group (P＜0.05). Transwell cell invasion experiment showed that the number of invasive cells in the miR-221 group was 23.42±3.62, which was significantly lower than that in the control group and the miR-221-NC group (P＜0.05). Transwell cell migration assay showed that the number of migrating cells in the miR-221 group was 34.86±5.32, which was significantly lower than that in the control group and the miR-221-NC group (P＜0.05). The relative level of PCNA, Cyclin D1 and MMP-9 in the miR-221 group was 0.26±0.03, 0.17±3.61 and 0.14±0.02, respectively, which was significantly lower than those in the control group and the miR-221-NC group (P＜0.05), while the relative level of Caspase-3 in the miR-221 group was 0.37±0.05, which was significantly higher than that in the control group and the miR-221-NC group (P＜0.05). Luciferase reporter assay showed that the activity of luciferase in Wnt wild type plasmid was significantly inhibited by miR-221 (P＜0.05). CONCLUSION: miR-221 can inhibit the proliferation, migration and invasion of BMNC, moreover can promote cell apoptosis, which may be related with the inhibition of Wnt/ß- catenin signaling pathway.

Cepharanthine hydrochloride reverses P­glycoprotein-mediated multidrug resistance in human ovarian carcinoma A2780/Taxol cells by inhibiting the PI3K/Akt signaling pathway.

Ovarian cancer has the highest mortality rate among gynecologic malignant tumors. The major obstacle to treatment success is multidrug resistance (MDR) to chemotherapy drugs. Cepharanthine hydrochloride (CH), a natural alkaloid-derived compound, has shown MDR reversal potency in several tumor cell lines; however, the molecular mechanism is not entirely known. In the present study, we assessed whether CH sensitized malignant cells to chemotherapy drugs in ovarian cancer and explored the relevant mechanism. We found that CH reduced the IC50 value of paclitaxel and increased intracellular rhodamine-123 accumulation in human ovarian cancer A2780/Taxol cells in a concentration-dependent manner. Reverse transcription polymerase chain reaction and western blot assay demonstrated that CH inhibited MDR1 expression as indicated by reduced mRNA and protein levels in A2780/Taxol cells. In addition, the inhibitory effect was strengthened after CH was combined with the specific PI3K/Akt signaling pathway inhibitor LY294002. Furthermore, p­Akt expression decreased gradually with the concentration of CH (2, 4 and 8 µM). Taken together, these findings indicated that CH reversed P­glycoprotein-mediated MDR in A2780/Taxol cells by inhibiting the PI3K/Akt signaling pathway.

Influence of electromagnetic pulse on the offspring sex ratio of male BALB/c mice.

Public concern is growing about the exposure to electromagnetic fields (EMF) and its effect on male reproductive health. Detrimental effect of EMF exposure on sex hormones, reproductive performance and sex-ratio was reported. The present study was designed to clarify whether paternal exposure to electromagnetic pulse (EMP) affects offspring sex ratio in mice. 50 male BALB/c mice aged 5-6 weeks were exposed to EMP daily for 2 weeks before mated with non-exposed females at 0d, 7d, 14d, 21d and 28d after exposure. Sex hormones including total testosterone, LH, FSH, and GnRH were detected using radioimmunoassay. The sex ratio was examined by PCR and agarose gel electrophoresis. The results of D0, D21 and D28 showed significant increases compared with sham-exposed groups. The serum testosterone increased significantly in D0, D14, D21, and D28 compared with sham-exposed groups (p<0.05). Overall, this study suggested that EMP exposure may lead to the disturbance of reproductive hormone levels and affect the offspring sex ratio.

Sesquiterpenes from the fruits of Illicium jiadifengpi and their anti-hepatitis B virus activities.

Two new compounds, 2S-hydroxyl-jiadifenolide (1) and jiadifenlactone acid (2), and eight known compounds were isolated from the fruits of Illicium jiadifengpi. Their structures were analysed using several spectroscopic techniques, including 1D-, 2D-NMR and HR-ESI-MS experiments. The anti-hepatitis B virus (HBV) activities of the isolates were evaluated via HBV transfection of the Hep G2.2.15 cell line. The inhibitory rates of the most active compounds, compounds 4 and 5, on the HBeAg and HBsAg expression were 28.85±3.15% and 17.53±1.81% and 37.93±2.74% and 23.47±9.52% at concentrations of 64.94µM and 61.35µM, respectively.

Arterial infusion of 5-fluorouracil combined with subcutaneous injection of pegylated interferon alpha-2b in treating unresectable hepatocellular carcinoma with portal vein tumor thrombus.

This study aimed to evaluate the efficacy of arterial infusion of 5-fluorouracil (5-FU) combined with subcutaneous injection of pegylated interferon alpha-2b (PEG-IFNα-2b) on unresectable HCC with PVTT and to determine the potential survival benefits of patients from this therapy. From January 2007 to August 2009, HCC patients with PVTT were assigned to undergo arterial infusion of 5-FU with subcutaneous injection of PEG-IFNα-2b or palliative therapy. A total of 105 HCC patients (94 males and 11 females) aged 24-78 years with portal vein thrombosis were randomly assigned to 5-FU/PEG-IFNα-2b group (n = 55) or control group (n = 50). The median progression-free survival (PFS) was 10 months, and the median overall survival (OS) was 12 months for all patients. The 6- and 12-month OS rates were significantly higher in 5-FU/PEG-IFNα-2b group than in control group (81.04 vs. 22.72%, 48.03 vs. 0%, both P < 0.05). The median OS and PFS were significantly longer in 5-FU/PEG-IFNα-2b group than in control group (14.7 vs. 4.5 months, 12.5 vs. 9 months, both P < 0.05). For unresectable HCC with PVTT, arterial infusion of 5-FU combined with subcutaneous injection of PEG-IFNα-2b reduces tumor size obviously, prolongs patient survival, and causes no severe adverse events.

Anti-hepatitis B virus activities and absolute configurations of sesquiterpenoid glycosides from Phyllanthus emblica.

During the process exploring anti-viral compounds from Phyllanthus species, eight new highly oxygenated bisabolane sesquiterpenoid glycoside phyllaemblicins G1­G8 (1­8) were isolated from Phyllanthus emblica, along with three known compounds, phyllaemblicin F (9), phyllaemblic acid (10) and glochicoccin D (11). Phyllaemblicin G2 (2), bearing a tricyclo [3.1.1.1] oxygen bridge ring system, is an unusual sesquiterpenoid glycoside, while phyllaemblicins G6­G8 (6­8) are dimeric sesquiterpenoid glycosides with two norbisabolane units connecting through a disaccharide. All the structures were elucidated by the extensive analysis of HRMS and NMR data. The relative configuration of phyllaemblicin G2 was constructed based on heteronuclear coupling constants measurement, and the absolute configurations for all new compounds were established by calculated electronic circular dichroism (ECD) using time dependent density functional theory. The sesquiterpenoid glycoside dimers 6­9 displayed potential anti-hepatitis B virus (HBV) activities, especially for the new compound 6 with IC50 of 8.53 ± 0.97 and 5.68 ± 1.75 µM towards the HBV surface antigen (HBsAg) and HBV excreted antigen (HBeAg) secretion, respectively.

A new sesquiterpene lactone from the fruits of Illicium henryi.

AIM: To study the chemical constituents of the fruits of Illicium henryi. METHOD: Chromatographic separations on silica gel, Sephadex LH-20 gel and MCI gel were used to isolate the compounds. The structures were elucidated based on extensive spectroscopic data analyses. RESULTS: Seven compounds were obtained and their structures were identified as 10-benzoyl-cycloparvifloralone (1), cycloparvifloralone (2), 2α-hydroxycycloparviforalone (3), henrylactone B (4), merrillianone (5), henrylactone C (6) and 7, 14-ortholactone- 3-hydroxyfloridanolide (7). CONCLUSION: Compound 1 is a new sesquiterpene lactone. The tested compounds showed weak anti-HBV activities on HBV surface antigen (HBsAg) secretion and HBV e antigen (HBeAg) secretion using Hep G2.2.15 cell line.

Inhibition of Wnt signaling induces cell apoptosis and suppresses cell proliferation in cholangiocarcinoma cells.

The aim of the present study was to explore possible gene therapy for hilar cholangiocarcinoma by detecting the activation of the Wnt signaling pathway in 4 cholangiocarcinoma cell lines and inhibiting its expression by RNA interference (RNAi) targeting key factors of this pathway. The expression levels of the Wnt pathway-related factors, Wnt2, Wnt3, ß-catenin and transcription factor 4, and its target genes, c-myc and cyclin D1, in 4 cholangiocarcinoma cell lines were detected by RT-PCR, western blotting and immunofluorescence microscopy. After transfection of siRNAs targeting Wnt2 and ß-catenin into FRH0201 cells, the expression of the Wnt pathway-related factors and its target genes was again detected, and the cell cycle distribution, apoptosis and proliferation were analyzed by flow cytometry and MTT assay. Activation of the Wnt pathway and the expression of its target genes were detected in all 4 cell lines at various levels. After siRNA transfection, the expression of the target genes in the FRH0201 cells was significantly downregulated. In addition, the Wnt pathway was blocked, cell apoptosis was enhanced and cell proliferation was suppressed. In conclusion, the Wnt signaling pathway is activated in cholangiocarcinoma cells. RNAi technology targeting Wnt2 and ß-catenin may be a possible gene therapy for hilar cholangiocarcinoma.

Effects of electromagnetic pulse on polydactyly of mouse fetuses.

There is an increasing public concern regarding potential health impacts from electromagnetic radiation exposure. Embryonic development is sensitive to the external environment, and limb development is vital for life quality. To determine the effects of electromagnetic pulse (EMP) on polydactyly of mouse fetuses, pregnant mice were sham-exposed or exposed to EMP (400 kV/m with 400 pulses) from Days 7 to 10 of pregnancy (Day 0 = day of detection of vaginal plug). As a positive control, mice were treated with 5-bromodeoxyuridine on Days 9 and 10. On Days 11 or 18, the fetuses were isolated. Compared with the sham-exposed group, the group exposed to EMP had increased rates of polydactyly fetuses (5.1% vs. 0.6%, P < 0.05) and abnormal gene expression (22.2% vs. 2.8%, P < 0.05). Ectopic expression of Fgf4 was detected in the apical ectodermal ridge, whereas overexpression and ectopic expression of Shh were detected in the zone of polarizing activity of limbs in the EMP-exposed group and in the positive control group. However, expression of Gli3 decreased in mesenchyme cells in those two groups. The percentages of programmed cell death of limbs in EMP-exposed and positive control group were decreased (3.57% and 2.94%, respectively, P < 0.05, compared with 7.76% in sham-exposed group). In conclusion, polydactyly induced by EMP was accompanied by abnormal expression of the above-mentioned genes and decreased percentage of programmed cell death during limb development.

Cytokine-induced killer cells co-cultured with complete tumor antigen-loaded dendritic cells, have enhanced selective cytotoxicity on carboplatin-resistant retinoblastoma cells.

Retinoblastoma (RB) is a challenging disease that affects mostly young children. Chemical therapy has been shown to have limitations during clinical practice, principally because of the ability of RB to become resistant to the treatment. Nevertheless, chemotherapy is still the main treatment for RB, and immunotherapy has become a promising treatment for most solid tumors with fewer side effects than traditional therapies. In this study, we explored the antitumor effects of cytokine-induced killer (CIK) cells co-cultured with dendritic cells (DCs) pulsed with complete tumor antigens (DC-Ag). Cytotoxicity and specificity were evaluated on an RB cell line (RB-Y79), on a human normal retina cell line (hTERT-RPE1) and a carboplatin-resistant RB cell line. Our results showed that CIK differentiation and cytotoxicity were enhanced by co-culturing CIKs with DC-Ag. Moreover, the co-culture improved the CIK proliferation rate by increasing IL-6 and decreasing IL-10 levels in the culture medium. Furthermore, the use of DC-Ag-CIK cells had little effect on normal retinal cells but high cytotoxicity on RB cells even on carboplatin-resistant retinoblastoma cells. This is the first study showing that DC cells pulsed with the complete tumor antigen improve proliferation, differentiation and cytotoxic activity of CIKs specific not only for RB but also for the chemotherapy-resistant form of the malady. Thus highly efficient immunotherapy based on DC-Ag-CIK cells may be a potential effective and safe mean of treating RB especially to patients where traditional chemical therapy has failed.