Anemoside B4 prevents chronic obstructive pulmonary disease through alleviating cigarette smoke-induced inflammatory response and airway epithelial hyperplasia.

BACKGROUND: Cigarette smoke (CS) is one of the major risk factors for chronic obstructive pulmonary disease (COPD) and increases the risk of lung cancer (LC). Anemoside B4 (B4) is the main bioactive ingredient in Pulsatilla chinensis (P. chinensis), a traditional medicinal herb for various diseases. It has a wide range of anti-inflammatory, anti-oxidation and anti-cancer activities. However, in recent years, there is no relevant literature report on the therapeutic effect of B4 on COPD, and the anti-inflammatory and inhibitory effects of anemoside B4 on basal cell hyperplasia in CS-induced COPD have not been clearly established. PURPOSE: In the present study, we investigated whether anemoside B4 could alleviate CS or cigarette smoke extract (CSE) induced inflammation of COPD and further prevent basal cell hyperplasia, hoping to find its possible mechanism. METHODS: In this study, a COPD mouse model was established in C57BL mice by CS exposure 3 months. Bronchial pathology and basal cell hyperplasia were observed by HE staining and immunostaining. The contents of glutathione peroxidase catalase (GSH-PX), malondialdehyde (MDA) and superoxide dismutase (MPO) were determined by GSH-PX, MDA and SOD assay kits, respectively. 16HBE cells were cultured with 5% CSE with or without treatment with B4 (1, 10, 100 µM) or DEX (20 µM) in vitro. Cell viability was assessed by a cell counting kit 8 (CCK-8). Reactive oxygen species (ROS) generation was tested by DCFH-DA. Moreover, anti-inflammatory mechanism of anemoside B4 was further determined by pro-inflammatory cytokines production using RT-PCR. Protein expression levels of MAPK/AP-1/TGF-ß signaling pathway were measured by western blot. RESULTS: Anemoside B4 improved the lung function of mice, relieved lung inflammation and reduced the MDA, MPO and GSH-Px in the plasma. At the same time, B4 repressed the oxidative stress response and played a role in balancing the levels of protease and anti-protease. During the process of bronchial basal cell hyperplasia, B4 alleviated the degree of cell hyperplasia, and prevented further deterioration of hyperplasia through increased P53 and inhibited FHIT protein. In addition, B4 reduced ROS levels in human bronchial epithelial cells stimulated by CSE in vitro study. Meanwhile, B4 treatment also significantly attenuated increased IL-1ß, TGF-ß, IL-8 and TNF-α from CSE treated human bronchial epithelial cells. The expression of p-P38, AP-1(c-fos, and c-Jun), TGF-ß proteins in MAPK/AP-1/TGF-ß signaling pathway were decreased and the signal cascade reaction was blocked. CONCLUSION: Anemoside B4 protects against CS-induced COPD. These findings indicated that B4 may have therapeutic potential for the prevention and treatment of COPD.

Anticancer effect of myristicin on hepatic carcinoma and related molecular mechanism.

CONTEXT: Myristicin is a natural active compound that has inflammatory, antimicrobial and anti-proliferative properties. Yet, its effect on hepatic carcinoma has not been investigated. OBJECTIVE: To explore the role and related molecular mechanism of myristicin in hepatic carcinoma in vitro. MATERIALS AND METHODS: Human hepatic carcinoma cell lines (Huh-7 and HCCLM3 cells) were treated with different concentrations of myristicin (0.5, 1 and 5 mM) for 24, 48 and 72 h. Then, (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) tetrazolium assay (MTT), flow cytometer (FCM) analysis and transwell assay were performed to determine cell proliferation, apoptosis and migration/invasion, respectively. Protein levels of B-cell lymphoma-2 (Bcl-2), Bcl-2 associated X (Bax), E-cadherin, N-cadherin and phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signalling pathway-related proteins were detected using Western blot assay. Gene expression was determined using quantitative real time-polymerase chain reaction (qRT-PCR). RESULTS: Myristicin inhibited cell proliferation and induced apoptosis in Huh-7 and HCCLM3 cells; suppressed cell migration and invasion ability, and increased E-cadherin expression and decreased N-cadherin expression, thereby inhibiting epithelial-mesenchymal transition (EMT). Finally, the findings indicated that myristicin decreased phosphorylated (p)-mTOR and p-AKT expression at the protein level. DISCUSSION AND CONCLUSIONS: Myristicin exerts an efficient therapeutic effect on hepatic carcinoma by suppressing PI3K/Akt/mTOR signalling pathway; thus, it may be used as a new potential drug for hepatic carcinoma treatment.

Roles of penA and mtrR gene mutations in in vitro-induced resistance of Neisseria gonorrhoeae to ceftriaxone / 中华皮肤科杂志

Objective To investigate the roles of penA and mtrR gene mutations in resistance of Neisseria gonorrhoeae to ceftriaxone.Methods Standard strains of Neisseria gonorrhoeae (ATCC-49226),clinical strains of Neisseria gonorrhoeae with high sensitivity to ceftriaxone (2012-4052 and 2012-15361) and clinical strains of Neisseria gonorrhoeae with reduced sensitivity to ceftriaxone (2012-5616) were treated with ceftriaxone at subinhibitory concentration (50％ MIC),so as to induce the resistance to ceftriaxone.DNA was extracted from the primary strains before the treatment and daughter strains resistant to ceftriaxone after the treatment,followed by the amplification and DNA sequencing of the penA and mtrR genes.Results For strains 2012-5616 and ATCC-49226,ceftriaxone-resistant strains with MIC ≥ 1 mg/L were obtained after 26 and 28 passages,respectively.For strains 2012-4052 and 2012-15361,ceftriaxone-resistant strains with MIC ≥ 0.5 mg/L were obtained after 22 and 36 passages,respectively.Sequence analysis of the penA gene revealed that A501T and G542S mutations were identified in the induced resistant ATCC-49226 strains,but no new mutations were observed in the other 3 strains.All the 4 mutant strains showed penicillin-binding protein 2 (PBP2) of gene sequence type ⅩⅧ and no mosaic structure of the penA gene was found in the strains.Sequence analysis of the mtrR gene showed that the A39T mutation was found in the 2012-5616 and ATCC-49226 strains before and after the induction,as well as in the coding region of the mtrR gene in the induced resistant 2012-4052 strains.Conclusion The A501T and G542S mutations in the penA gene and A39T mutation in the mtrR gene may play a role in the resistance of Neisseria gonorrhoeae to ceftriaxone.

Relationship between B-tupe natriuretic peptide and preeclampsia of hypertensive disorder complicating pregnancy as well as its significance / 中国综合临床

Objective To investigate and evaluate the correlation between brain natriuretic peptide (BNP) and gestational hypertension and preeclampsia of hypertensive disorder complicating pregnancy (HDCP).Methods Fifty cases with HDCP and 46 cases with mild and 83 cases with sever stage preeclampsia were selected as our subjects.And 33 cases with regular pregnancy and 31 with irregular pregnancy were served as control group.Plasma brain natriuretic peptide,urinary protein quantity(UBQ),24-hour urinary protein assay (UPA) were measured.The correlations of brain natriuretic peptide and UBQ,UPA,systolic pressure (SP),diastolic pressure (DP) were analyzed.Results The levels of brain natriuretic peptide in the group with gestational hypertension and mild,severe preeclampsia groups were (48.54± 18.27),(79.46± 32.18) and (292.24±213.08) ng/L,higher than that in normal pregnancy and non pregnant group ((27.84± 14.58) and (20.63± 8.28) n/L;F =49.583,P＜0.05).While no significant difference exists between normal pregnancy group and non pregnant group.Grouped on the median values (199) of brain natriuretic peptide of the severe preeclampsia group,the levels of 24-hour UPA,systolic pressure and diastolic pressure were (5.46±2.68) g,(174.55± 13.58) mmHg,(113.74±9.91) mmHg in patients with brain natriuretic peptide ≥ 199 ng/L(n=42),significant higher than those in patients with brain natriuretic peptide ＜ 199 ng/L(n =41;(4.34± 1.95)g,(165.31±11.12) mmHg,(106.05±8.02) mmHg;t=2.603,3.396,2.308;P=＞0.010,0.001,0.024).The levels of 24-hour UPA,systolic pressure and diastolic pressure of patients with brain natriuretic peptide ≥ 86ng/L(n=20) in mild preeclampsia were (1.68±0.27) g,(163.69±8.29) mmHg,(105.45±6.71) mmHg,significant higher than those in patients with brain natriuretic peptide ＜ 86 ng/L (n =26;(1.16 ± 0.31) g,(152.90±7.32) mmHg,(99.19 ± 5.25) mmHg;t =3.180,2.508,2.32;P =0.010,0.016,0.025).Brain natriuretic peptide was closely correlated with UPA,systolic pressure and diastolic pressure in hypertensive disorder complicating pregnancy (HDCP) (r =0.29,0.30;P ＜ 0.01).Brain natriuretic peptide was closely correlated with UPA systolic pressure and diastolic pressure in mild preeclampsia (r =0.39,0.37,0.40;P ＜0.01).And correlation efficacy of brain natriuretic peptide with UPA,systolic pressure and diastolic pressure were 0.44,0.42 and 0.53 (P＜0.01).Conclusion The level of brain natriuretic peptide is closely associated with the severity of gestational hypertension and preeclampsia of hypertensive disorder complicating pregnancy.Correlation of brain natriuretic peptide to the severity of gestational hypertension and preeclampsia is independent of urinary protein and hypertension.Brain natriuretic peptide is an important indicator for the severity of gestational hypertension and preeclampsia of hypertensive disorder complicating pregnancy.

Association of Ureaplasma urealyticum biovar and genotype with mucopurulent cervicitis / 中华皮肤科杂志

Objective To study the association of U. urealyticum biovar and genotype with nongonococcal, nonchlamydial mucopurulent cervicitis. Methods The study population consisted of two groups: patient group (226 female patients with nongonococcal, nonchlamydial mucopurulent cervicitis) and control group (118 healthy women). The biovar and genotype of U. urealyticum were identified in specimens positive for U. urealyticum culture by using PCR-single-strand conformation polymorphism (PCR-SSCP) analysis. Results The most common genotype in both groups was mba 3/14 in biovar 1 with the detection rate being 30.98%(57/184) in patients with mucopurulent cervicitis and 43.42% (33/76) in the controls. A significant difference was observed in the prevalence of genotype 2B in biovar 2 between the patients and controls [16.30% (30/184) vs 6.58% (5/76), χ2 = 4.367, P= 0.037). The genotype 1, 3/14 and 6 in biovar 1 predominated in the controls with their total prevalence being 81.58%. Conclusion The genotype 2B in biovar 2 of U. urealyticum may be associated with nongonococcal, nonchlamydial mucopurulent cervicitis among female patients attending an STD clinic.

Effects of simvastatin on the mouse model of sclerotic skin / 中华皮肤科杂志

Objective To study the effect of simvastatin on the mouse model of sclerotic skin. Methods A total of 44 mice were divided into two groups, i.e., early administration group (n=24) and post-induction administration group (n=20), and the former was classified into three subgroups, including negative group, model group and simvastatin-treated group, and the latter into two groups, namely blank control group, simvastatin-treated group. The mouse model of sclerotic skin was established by local injec-tions of bleomycin in the back of BALB/c mice. Simvastatin was administered by gavage at a dose of 5 μg per kilogram body weight per day for 4 weeks to mice at the same time when bleomycin was injected in the early group or after 4-week bleomycin injection in the post-induction group. Skin sections were prepared 24 hours after the last administration of simvastatin for histopathological examination and measurement of derma l thickness with HE staining, determination of hydroxyproline content via colorimetry, and mRNA expression of procollagen α1 (Ⅰ) by RT-PCR. Results In the early administration group, a significant increment was observed in the diameter of dermal collagen, skin thickness, and hydroxyproline content in model group compared with the negative control group (all P <0.01), whereas decreased dermal thickness, hydroxyproline content and mRNA expression of procollagen α1(Ⅰ) were noticed in simvastatin-treated group in comparison with the model group (P<0.05, 0.01 and 0.05, respectively). No obvious improvement was achieved in dermal thickness or hydroxyproline content in simvastatin-treated group compared with blank control group (both P0.05), but the mRNA expression of procollagen α1 (Ⅰ) was inhibited in the former group (P<0.05). Conclusion Skin sclerosis is relieved significantly by administration of simvastatin at the induction of scle- rosis but not by that after the induction of sclerotic skin.

Relationship between Mycoplasma genitalium and mueopurulent cervicitis / 中华皮肤科杂志

Objective To study the relationship between M. Genitalium and nongonococcal, non-chlamydial mucopurulent cervicitis. Methods A total of 226 patients with nongonococcal, nonchlamydial mu copurulent cervicitis and 118 healthy female volunteers were recruited in this study. Cervical samples were collected and M.genitalium was detected by PCR amplification of 16s rRNA and Pa genes. Information about socio-demographic characteristics, medical history, and sexual behaviors was gathered by questionnaire from both populations. Results The prevalence of M. Genitalium infection was 11.06% (25/226) in patients with mucpurulent cervicitis, 0,85% (1/118) in the healthy controls; the difference was significant between the two groups (x2 = 11.58, P < 0.001). Single variant analysis on the 226 patients showed that the preva-lence of M. Genitalium infection was 27.78%, 16.36%, 18.28% and 14.12% in patients with ectopic pregnancy history, cervical inflammation, pelvic organ tenderness, 10 or more polymorphonuclear leukocytes (PMNs)per oil immersion field in cervical discharge, respectively, significantly higher than that in patients without ectopic pregnancy history, cervical inflammation or pelvic organ tenderness, and those with less than 10 PMNs per oil immersion field in cervical discharge (9.62%, 6.03%, 6.02% and 1.79%, all P < 0.05). M.genitalium infection was also related to multiple sex partners and the presence of mucopurulent secretion in cervix (P < 0.001). Conclusion The prevalence of M.genitalium infection is higher in patients with non-gonococcal, nonchlamydial mucopurulent cervicitis attending STD clinic than that in normal population.

Amino acid substitution or insertion patterns in penicillin-binding protein 2 in Neisseria gonorrhoeae isolates with reduced susceptibility to ceftriaxone / 中华皮肤科杂志

Objective To investigate the amino acid patterns in penicillin-binding protein 2(PBP2)in Neisseria gonorrhoeae isolates with reduced susceptibility to ceftriaxonc.and the relationship between the amino acid patterns and reduced ceftriaxone susceptibility.Methods DNA was extracted from 13 clinical isolates of N.gonorrhoeae.including 11 strains with decreased susceptibility to ceftriaxone and 2 sensitive isolates.The full-length penA gene encoding the penicillin-binding protein 2 was amplified and sequenced.BLASTn and BLASTx programs were used to assess the insertion and substitution patterns of nucleotides in penA gene and of amino acids in PBP2,respectively.Results BLASTn analysis revealed insertion or substitution of 18-38 nucleotides in the penA gene of gonococcal isolates with reduced ceftriaxone susceptibility.As shown by BLASTX analysis.there were five patterns of amino acid substitution or insertion in PBP2 of the 11 isolates with reduced ceftriaxone susceptibility.However.mosaic structure of PBP2 was not found in any of these isolates.Conclusion Mosaic PBP2 seems not to be the major factor contributing to the decrease in susceptibility of N.gonorrhoeae to ceftriaxone.

Correlation of Serum Cystatin C,Urinary Cystatin C and Renal Pathology in Primary Glomerulonephritis Patients of Different Patological Types / 医学研究杂志

Objective To investigate the variability of serum Cystatin C concentration and urinary Cystatin C concentration in primary glomerulonephritis patients of different patological types,and to study the correlation with renal pathological changes. Methods Urinary Cystatin C was detected before renal biopsy.The two indexes were processed by correlation analysis with glomcrulus scores and renal tubulointerstitial scores respectively.Results The differences of Serum Cystatin C concentration and urinary Cystatin C concentration in IgA nephropathy(IgAN) group,focal segmental glomerulosclerosis group and membranous nephropathy group existed statistical significance as compared to normal control (P