OBJECTIVETo investigate the potential protective effect of the mitochondria-targeted antioxidant Mitoquinone (MitoQ) on post-thaw human sperm.

METHODSSemen samples were collected from 60 normal fertile men, each divided into six parts of equal volume to be incubated at 37 °C in normal saline (G0, control) or in the extender with 2 nmol/L (G1), 20 nmol/L (G2), 200 nmol/L (G3), 2 µmol/L (G4), and 20 µmol/L of MitoQ (G5). After one hour of incubation, the samples were subjected to computer-assisted semen analysis (CASA) for sperm motility, flow cytometry for reactive oxygen species (ROS), thiobarbituric acid assay for the concentration of malondialdehyde (MDA), and MitoTracker fluorescent staining and flow cytometry for the sperm mitochondrial membrane potential (MMP). Then, the semen were cryopreserved with none (B0), 200 nmol/L (B1), and 2 µmol/L of MitoQ (B2), followed by detection of the changes in the ROS, MDA, and MMP of the post-thaw sperm.

RESULTSThe percentage of progressively motile sperm and total rate of sperm motility were significantly higher in G3 ([30.8 ± 10.2]% and [70.6 ± 9.0]%) and G4 ([32.7 ± 13.5]% and [70.3 ± 11.9]%) than in G0 ([17.6 ± 5.0]% and [54.9 ± 11.5]%) (P < 0.05). The level of ROS dropped markedly with the increased concentration of MitoQ, 86.5 ± 31.6 in G3, 93.6 ± 42.0 in G4, and 45.1 ± 15.0 in G5, as compared with 160.8 ± 39.7 in G0 (P < 0.05). The content of MDA was remarkably lower in G3 ([0.9 ± 0.5] µmol/mg) and G4 ([0.9 ± 0.5] µmol/mg) than in G0 ([1.9 ± 1.1] µmol/mg) (P < 0.05), but not in G5 ([1.7 ± 0.7] µmol/mg), which was even higher than in G3 and G4 (P < 0.05). The MMP showed a significant reduction in G5 (1156 ± 216) in comparison with G0 (1701 ± 251) (P < 0.05) but exhibited no remarkable difference between G0 and G1 (1810 ± 298), G2 (1995 ± 437), G3 (1950 ± 334), or G4 (1582 ± 314). The percentage of progressively motile sperm and total rate of sperm motility after freezing-thawing were significantly decreased as compared with those of the fresh semen (P < 0.01), but both were remarkably higher in B1 ([3.2 ± 2.3]% and [ 43.0 ± 9.5]%) than in B0 ([0.8 ± 0.6]% and [26.5 ± 11.4]%) (P < 0.05). The ROS level was significantly lower in B1 and B2 than in B0 (34.6 ± 12. 3 and 37.0 ± 10.5 vs 56.9 ± 14.3, P < 0.05), and so was the MDA content ([1.4 ± 0.5] and [1.4 ± 0.6] µmol/mg vs [2.6 ± 1.0] µmol/mg, P < 0.05), but the MMP was markedly higher in B1 and B2 than in B0 (1010.0 ± 130.5 and 880.6 ± 128.6 vs 721.1 ± 24.8, P < 0.05).

CONCLUSIONAddition of MitoQ to the freezing extender at 200 nmol/L may effectively improve the quality of human sperm and MitoQ is a good protective addictive for human sperm cryopreservation.

Antioxidants ; Cryopreservation ; Humans ; Male ; Malondialdehyde ; analysis ; Membrane Potential, Mitochondrial ; Mitochondria ; Organophosphorus Compounds ; pharmacology ; Oxidative Stress ; drug effects ; Reactive Oxygen Species ; Semen ; Semen Analysis ; Semen Preservation ; Sperm Motility ; Spermatozoa ; drug effects ; Ubiquinone ; analogs & derivatives ; pharmacology

OBJECTIVETo provide application basis of the Forsythia suspensa by studying the difference of HPLC-FP of F. suspensa fructification (medicinal materials).

METHODComparative work was done on F. suspensa produced in different areas, on different parts of Forsythia suspensa, and on the pseudo preducts with methods of HPLC-FP.

RESULTDifferent FP characteristics were shown respectively by different samples, which were from different producing areas, from different parts, and the pseudo products including the fructification of Syringa reticulata var. and F. viridissimac.

CONCLUSIONThe FP can be used to distinguish the F. suspensa coming from different producing areas and different sources.

Chromatography, High Pressure Liquid ; Chromatography, Paper ; methods ; Drug Contamination ; Drugs, Chinese Herbal ; chemistry ; Forsythia ; chemistry ; classification ; Fruit ; chemistry ; Peptide Mapping ; Plants, Medicinal ; chemistry ; Species Specificity

Purpose To investigate the pathological features and prognostic value of tertiary lymphoid structure (TLS) formation in gastric cancer (GC) . Methods HE staining slides were reviewed to evaluate the TLS in 163 specimens from patients with GC in the Third Affiliated Hospital of Soochow University from 2006 to 2008. The validation cohort contained 63 randomly selected cases and immunohistochemical staining of MECA-79 was used to verify the accuracy of pathological assessment of TLS. Results TLS score and MECA-79 immunohistochemical staining showed significant correlation (P = 0. 002) and agreement (P = 0. 024) . The TLS was not significantly correlated to clinical pathological parameters. The patients with high level of TLS had better prognosis (P = 0. 025) with the mean survival time of 48. 54 months. In multivariate Cox regression analysis, TLS was an independent prognostic factor (P = 0. 031) . Conclusion The pathological evaluation of TLS is accurate. The formation of TLS is an important positive prognostic factor for GC patients.

OBJECTIVETo study the flavanoids extracted from onion on the blood-brain barrier (BBB) permeation, and their effects on primary cultured neuron cell proliferation and apoptosis of SD rats using ethanol reflux method.

METHODSThe brain microvascular endothelial cells (BMVECs) were first successfully primary cultured. Then rats BMVECs and astrocytes (ACs) were co-cultured to establish the in vitro BBB model. The flavanoids were extracted from onion using ethanol reflux method. The model was verified by transmission electron microscopy (TEM) and trans-epithelial electric resistance (TEER). The flavanoids permeability was tested using high performance liquid chromatography (HPLC). Meanwhile, rat neuron cells were cultured and exposed to H2O2 and flavanoids. Their effects on the cell proliferation and apoptosis were observed using MTT assay. The injury of neuron DNA was analyzed using single-cell gel electrophoresis (SCGE) and immunofluorescent assay.

RESULTSThe in vitro BBB model was successfully established by TEM and TEER. Results of HPLC proved flavanoids extracts could effectively permeate the BBB with the permeability of 60.58%. The extractive at 10 - 20 microg/mL showed obvious inhibition on the apoptosis of neuron cells induced by H2O2, and attenuated the injury of neuron DNA.

CONCLUSIONSThe flavanoids extracted from onion ethanol reflux method could effectively penetrate the BBB. They also showed obvious inhibition on the H2O2 induced neuron cell apoptosis and DNA injury.

Animals ; Animals, Newborn ; Apoptosis ; Astrocytes ; cytology ; drug effects ; Blood-Brain Barrier ; drug effects ; Cells, Cultured ; DNA Damage ; Endothelial Cells ; cytology ; drug effects ; Flavonoids ; pharmacology ; Hydrogen Peroxide ; Neurons ; cytology ; drug effects ; Neuroprotective Agents ; pharmacology ; Onions ; chemistry ; Rats

OBJECTIVETo analyse the cartilage erosion related blood biochemical and immune factors in rheumatoid arthritis (RA) and to explore the special influences of Chinese medicine (CM) and Western medicine (WM) on these factors.

METHODSThree hundred and ninety-seven patients, with confirmed diagnosis of active RA, were randomly assigned to the WM group (194 patients) and the CM group (203 patients). The WM applied covered non-steroid anti-inflammatory agents and slow acting medicine; and the CM given included basic remedy and syndrome differentiating medication. Related blood biochemical and immunological indexes were determined before and after treatment to screen out the cartilage erosion related factors and to compare the influence of CM and WM on them.

RESULTSPatients' peripheral red blood cell (RBC) and platelet (PLT) count were changed closely along with their degree of cartilage erosion. RBC count increased in the CM group and PLT count lowered in the WM group after treatment, all showed statistical significance; comparison of the two indexes between the two groups showed that statistical difference presented in RBC but not in PLT count.

CONCLUSIONBoth WM and CM can ameliorate the cartilage erosive factor in RA, but they are acting in different ways.

Adult ; Antirheumatic Agents ; therapeutic use ; Arthritis, Rheumatoid ; blood ; drug therapy ; Cartilage, Articular ; drug effects ; pathology ; Drugs, Chinese Herbal ; therapeutic use ; Erythrocyte Count ; Female ; Humans ; Male ; Methotrexate ; therapeutic use ; Middle Aged ; Phytotherapy ; Platelet Count ; Prednisone ; therapeutic use ; Young Adult

Objective:To study the predictive value of the amplitude-integrated electroencephalography (aEEG) within 6 hours and 3 days after birth and magnetic resonance imaging(MRI) on the adverse neurobehavioral development of asphyxiated preterm infants at the correction age of 6 months.Methods:From December 2017 to June 2019, 50 asphyxiated preterm infants who were delivered at the obstetrical department transferred to the division of neonatology in the Third Affiliated Hospital of Anhui Medical University were monitored by aEEG within 6 hours after birth, then once a day for at least 4 h. MRI was administered at 40 weeks of corrected age, neuromotor developmental function of the infants was assessed by the Geisel developmental diagnostic scale at 6 months of corrected age, then the infants were divided into good prognosis group and poor prognosis group according to the assessment results. SPSS 19.0 software was used for statistical analysis.The software of SPSS 19.0 was used to analyze the data.Independent sample t-test and χ 2 test were used to analyze the difference between the two groups.The relationship between aEEG grading and MRI, and their predictive value for adverse neurobehavioral development were analyzed at 6 months of corrected age. Results:The degree of white matter damage( H=24.896) and intracranical hemorrhage( H=29.245) of premature infants with different aEEG clinical grades were different (both P<0.01) on MRI. The sensitivity of aEEG within 6 hours and 3 days after birth on predicting poor prognosis was 96.2% and 97.8%, the specificity was 56.2% and 62.5%, the negative predictive value was 98.2% and 99.0%, the positive predictive value was 37.8% and 52.3%, the correct index was 52.4% and 60.3%, respectively. The aEEG was combined with MRI, the sensitivity (90.0%, 97.0%), specificity (89.0%, 99.0%), negative predictive value (99.2%, 99.5%), positive predictive value (80.6%, 88.5%), and correct index (79%, 96%) were all improved. Conclusion:The combination of aEEG grading and MRI can improve the prognostic value on neurodevelopmental prognosis, and provide a better evaluation basis for clinical follow-up and intervention of asphyxiated premature infants with brain injury.

Background The diagnosis and treatment of fungal keratitis are knotty.There is no quantitative method to identify the disease and judge the therapeutic effect of the antifungal agent.Studies have determined that serum (1,3-) β-D-glucan level can sensitively and specifically reflect the state of systemic mycotic-causing diseases.However,whether (1,3-) β-D-glucan level in tear can monitor and diagnose mycotic keratitis is unclear.Objective Purpose of this study was to investigate the change of tear (1,3-) β-D-glucan level following the administration of antifungal drug in fungal keratitis patients,and evaluate the diagnosis and monitor value of (1,3-) β-D-glucan in tears for fungal keratitis.Methods Sixty patients who were diagnosed as fungal keratitis by fungal culture were analyzed in Affiliated Hospital of Qingdao University Medical College from July 2010 to May 2012.The patients received the topical administration of antifungal drug for 28 days.Thirty healthy volunteers without eye disease served as normal controls.The tear of 50 μl was collected from each subject for the detection of (1,3)-β-D-glucan before the therapy,7,14,28 days after therapy and 7 days,14 days after the drugs were stopped,respectively.The dynamic changes of (1,3-) β-D-glucan levels in tears were evaluated and compared with the manifestation of the lesions under the laser scanning confocal microscope.The patients without hyphal by the laser scanning confocal microscopy and tear (1,3-)β-D-glucan level less than 20 ng/L were subsequently treated for another 7 days,and the following-up duration was 2 months.The informed consent was obtained before any medical examination was performed from each subject.Results (1,3-)β-D-glucan level in tears (Log value) was (6.37 ±0.48)ng/L in the patient group,and was significantly higher than (2.00±0.31) ng/L in the normal control group (t =2.89,P＜0.01).The lesion was smaller with the gradually clear border,and the number of mycelia was decreased under the laser scanning confocal microscope 7 days after treatment.(1,3-) β-D-glucan level in tears was gradually declined in a time-dependent manner after treatment.The (1,3)-β-D-glucan level in tears (Log) was (5.19 ± 0.42),(4.16 ± 0.33),(2.99 ±0.42),(2.91 ±0.39),(2.80±0.40) ng/L 7,14,28 days after treatment,and 7 days,14 days after the drugs were stopped,respectively,with a statistically significant difference in comparison with (6.37±0.48)ng/L before treatment (P＜0.01).(1,3)-β-D-gluean level in tears remained a lower level till the end of follow-up,and no recurrence of lesion was found in the patient group.Conclusions Detecting (1,3)-β-D-glucan level in tears is of good diagnosis and monitor value in the evaluation of fungal keratitis.

The deubiquitinating enzyme ubiquitin specific peptidase 15 (USP15) is regarded as a regulator of TGFβ signaling pathway. This process depends on Smad7, the inhibitory factor of the TGFβ signal, and type I TGFβ receptor (TβR-I), one of the receptors of TGFβ. The expression level of USP15 seems to play vital roles in the pathogenesis of many neoplasms, but so far there has been no report about USP15 in psoriasis. In this study, immunohistochemical staining of USP15, TβR-I and Smad7 was performed in 30 paraffin-embedded psoriasis specimens and 10 normal specimens to investigate the expression of USP15, TβR-I and Smad7 in psoriasis and to explore the relevance among them. And USP15 small interfering RNA (USP15 siRNA) was used to transfect Hacat cells to detect the mRNA expression of TβR-I and Smad7. Of 30 cases of psoriasis in active stage, 28, 24 and 26 cases were positive for USP15, TβR-I and Smad7 staining, respectively. The positive rates of USP15 and Smad7 were significantly higher in psoriasis specimens than in normal skin specimens (44.1%±26.0% vs. 6.1%±6.6%, 47.2%±27.1% vs. 6.6%±7.1%), and positive rate of TβR-I (20.3%±22.2%) in psoriasis was lower than that in normal skin specimens (46.7%±18.2%). There was a significant positive correlation between USP15 and Smad7 expression, and significant negative correlations between USP15 and TβR-expression, an I d between TβR- and Smad7 expression I in psoriasis. After transfection of USP15 siRNA in Hacat cells, the expression of TβR-mRNA was up I -regulated and that of Smad7 was down-regulated. It is concluded that USP15 may play a role in the pathogenesis of psoriasis through regulating the TβR-I/Smad7 pathway and there may be other cell signaling pathways interacting with USP15 to take part in the development of psoriasis.

OBJECTIVETo investigate the effect of beta-catenin on the activation of hepatic fibrosis by transforming growth factor-beta 1 (TGFbeta1).

METHODSThe recombinant expression plasmids pcDNA3.1(+)-beta-catenin and pEGFP-N1 were cotransfected into cultured HSC-T6 cells. The expression of smad3, beta-catenin and alpha-SMA, beta-catenin protein in TGFbeta1 treated HSC-T6 cells were detected by RT-PCR and Western-blot.

RESULTSThe expression of smad3 and beta-catenin in the co-transfected cells was higher than that in the untransfected cells (smad3 mRNA were 0.642 +/- 0.011, 0.501 +/- 0.021, 0.511 +/- 0.019, 0.356 +/- 0.017, respectively, F = 135.304, P < than 0.05. beta-catenin mRNA were 0.783 +/- 0.021, 0.543 +/- 0.033, 0.538 +/- 0.024, 0.212 +/- 0.019, respectively, F = 267.340, P < than 0.05. smad3 protein were 0.892 +/- 0.012, 0.124 +/- 0.011, 0.130 +/- 0.021, 0.003 +/- 0.001, F = 2823.813, P < l than 0.05. beta-catenin protein were 0.921 +/- 0.020, 0.210 +/- 0.010, 0.208 +/- 0.008, 0.002 +/- 0.001, respectively, F = 3440.982, P < than 0.05). The expression of beta-catenin and smad3 protein had a positive correlation with the level of alpha-SMA protein in cells (r = 0.901, P < than 0.01; r = 0.939, P < than 0.01).

CONCLUSIONSExpression of smad3/alpha-SMA/beta-catenin is increased in the cultured HSC-T6 cells transfected by beta-catenin gene, especially when the transfected cells are stimulated by TGFbeta1. Our data suggest that beta-catenin could aggravate hepatic fibrosis induced by TGFbeta1.

Actins ; metabolism ; Animals ; Blotting, Western ; Cell Line ; Green Fluorescent Proteins ; genetics ; Hepatic Stellate Cells ; metabolism ; Liver Cirrhosis, Experimental ; etiology ; metabolism ; pathology ; Plasmids ; genetics ; RNA, Messenger ; metabolism ; Rats ; Reverse Transcriptase Polymerase Chain Reaction ; Smad3 Protein ; genetics ; metabolism ; Transfection ; Transforming Growth Factor beta1 ; pharmacology ; beta Catenin ; genetics ; metabolism

BACKGROUNDHepatitis B virus infection is closely related to hepatocellular carcinoma (HCC). Cyclooxygenase-2 (COX-2) is overexpressed in HCC and considered to play a role in hepatic carcinogenesis. In this study, we analyzed the polymorphism of COX-2 promoter -899G/C in healthy controls, chronic hepatitis B (CHB) patients, liver cirrhosis patients, and hepatocellular carcinoma (HCC) patients, to investigate the relationship between COX-2 -899G/C polymorphism and the risk for hepatitis B-related liver cancer in a Chinese population from Gansu province.

METHODSPatients were divided into four groups: 300 patients with CHB, 300 patients with liver cirrhosis, 300 patients with HCC, and 300 healthy controls. The polymorphism of COX-2 -899G/C was detected by PCR-TaqMan probes. The results were analyzed by SPSS 17.0.

RESULTSThe COX-2 -899G/C genotypes were GG, GC, and CC. Frequencies in CHB were 87.00%, 12.67%, 0.33%; in liver cirrhosis were 85.33%, 14.00%, 0.67%; in HCC were 77.00%, 21.67%, 1.33%; and in healthy controls were 90.67%, 9.00%, 0.33%, respectively. COX-2 -899C carriers may have an increased risk for hepatitis B-related liver cancer. Compared with the frequency of GG genotype, there were significant differences in the frequency of GC genotype between HCC and healthy control groups (OR = 2.835, 95%CI: 1.751 - 4.589); HCC and CHB groups (OR = 1.933, 95%CI: 1.248 - 2.994); and HCC and liver cirrhosis groups (OR = 1.175, 95%CI: 1.119 - 2.628). Stratification analyses showed that COX-2 -899C allele carriers with a drinking history are more susceptible to develop HCC.

CONCLUSIONCOX-2 -899C genotype may increase the susceptibility of individuals to hepatitis B-related liver cancer in Gansu province, China.

Adult ; Aged ; Aged, 80 and over ; Asian Continental Ancestry Group ; Carcinoma, Hepatocellular ; etiology ; genetics ; Cyclooxygenase 2 ; genetics ; Female ; Genetic Predisposition to Disease ; Hepatitis B ; etiology ; genetics ; Humans ; Liver Neoplasms ; etiology ; genetics ; Male ; Middle Aged ; Polymorphism, Genetic ; genetics ; Promoter Regions, Genetic ; genetics