Baicalein suppresses the SOS response system of Staphylococcus aureus induced by ciprofloxacin.

Numerous antibiotics can induce an SOS repair system in bacteria that leads to antibiotic-resistant mutation of the bacterium. Therefore, searching for drugs that can prevent the SOS response and thus improve the long-term viability of some antibiotics is important. In this study, we aimed to detect the suppressive effects of baicalein on the SOS system and rifampin-resistant mutation in Staphylococcus aureus. We determined the reactive oxygen species (ROS) formation and intracellular ATP level in S. aureus with baicalein treatment to investigate the mechanisms involved in its effects on the SOS system. The results showed that baicalein was a potent inhibitor of the expression of the SOS genes RecA, LexA and SACOL1400. The rifampin-resistant mutation rate of S. aureus induced by ciprofloxacin was significantly reduced after treatment with baicalein. Treatment with baicalein led to a significant decrease in intracellular reactive oxygen species (ROS) formation and ATP level. Our findings indicate that baicalein may be an SOS-response inhibitor in S. aureus through inhibiting ROS formation and ATP production and may be used to prevent excessive mutation induced by antibiotics.

Green tea extract weakens the antibacterial effect of amoxicillin in methicillin-resistant Staphylococcus aureus infected mice.

Tea (Camellia sinensis) has been known for its modulation of resistance of methicillin-resistant Staphylococcus aureus (MRSA) to beta-lactam antibiotics in vitro. This study aimed to confirm the in vitro effect of green tea extracts with beta-lactams and to determine whether green tea extracts can reduce the minimum inhibitory concentrations (MICs) of amoxicillin in MRSA-infected mice. The catechins in the test tea that account for the reduced resistance to beta-lactams were quantitatively determined by high-performance liquid chromatography. The MICs of the ampicillin, cefazolin, amoxicillin, oxacillin, tea extract alone and tea extract in combination with beta-lactams were determined. Proportions of tea extracts and amoxicillin-tea extract combinations were administered to groups of mice enterally. The in vitro experiment showed that the MICs of four beta-lactams were greatly decreased in the presence of 0.25% tea extract. However, in an in vivo experiment, amoxicillin in combination with 5% tea extract conferred a higher ED(50) than that of antibiotic alone. Green tea extract, alone or in combination with amoxicillin, does not have protective benefits in MRSA-infected mice. This study concluded that tea extract weakened the antibacterial effect of amoxicillin in MRSA infected mice. Tea drinking is not recommended in combination with amoxicillin treatment.

FOXO3a is involved in the apoptosis of naked oocytes and oocytes of primordial follicles from neonatal rat ovaries.

Inhibition of the forkhead transcription factor, FOXO3a, can promote the transition from primordial to primary follicle and subsequent follicle development in mammalian ovaries. Stem cell factor (SCF) initiates anti-apoptotic signaling from its membrane receptor, c-kit, to Bcl-2 family members through PI3K/AKT in oocytes of primordial follicles. However, whether FOXO3a mediates the apoptosis of naked oocytes and oocytes of primordial follicles remains unknown. In the present study, oocytes from nests and primordial follicles from neonatal rat ovaries were cultured, and oocyte apoptosis was examined using the TUNEL technique. The pro-apoptotic action of FOXO3a and the potential signal transduction pathways were investigated using RT-PCR, Western blot, and immunocytochemistry. Culturing oocytes in the presence of SCF did not affect the level of total FOXO3a protein, but rapidly elevated the level of phosphorylated FOXO3a (indicating functional suppression). As phosphorylated FOXO3a increased, oocyte apoptosis was inhibited. The specific PI3K/Akt inhibitor, LY 294002, abolished the phosphorylation of FOXO3a and the anti-apoptotic action of SCF. SCF down-regulated the expression of p27KIP1 and pro-apoptotic factors such as Bim, Bad, and Bax, and this activity was reversed by LY 294002. SCF up-regulated the expression of MnSOD, which was also inhibited by LY 294002. However, SCF had no effect on Bcl-2 protein. These results suggest that FOXO3a is involved in oocyte apoptosis in the neonatal rat ovary, and the SCF-PI3K/Akt-FOXO3a signaling pathway mediates oocyte apoptosis and primordial follicle formation.

Incidence of extended-spectrum beta-lactamases and characterization of integrons in extended-spectrum beta-lactamase-producing Klebsiella pneumoniae isolated in Shantou, China.

This study is concerned with the level of antibiotic resistance of extended-spectrum beta-lactamase (ESBL)-producing Klebsiella pneumoniae, isolated in Shantou, China, and its mechanism. Seventy-four non-repetitive clinical isolates of K. pneumoniae producing ESBLs were isolated over a period of 2 years. Antibiotic susceptibility, carried out by Epsilometer test, showed that most of the isolates were multiresistant. Polymerase chain reaction showed that, among the several types of beta-lactamases, SHV was the most prevalent, TEM was the second most prevalent, and CTX-M was the least prevalent. Sixty-nine isolates were positive for integrase gene IntI1, but no IntI2 or IntI3 genes were found. The variable region of class 1 integrons were amplified and further identified by sequencing. Thirteen different gene cassettes and 11 different cassette combinations were detected. Dfr and aadA cassettes were predominant and cassette combinations dfrA12, orfF and aadA2 were most frequently found. No gene cassettes encoding ESBLs were found. Integrons were prevalent and played an important role in multidrug resistance in ESBL-producing K. pneumoniae.

Genome-wide expression profiling of the response to terbinafine in Candida albicans using a cDNA microarray analysis.

BACKGROUND: Candida albicans is the most frequently seen opportunistic human fungal pathogen. Terbinafine is an allylamine antifungal agent that has been proven to have high clinical efficacy in the therapy of fungal infections, the mechanism of action of terbinafine involves the specific inhibition of fungal squalene epoxidase, resulting in ergosterol deficiency and accumulation of intracellular squalene. We used cDNA microarray analysis technology to monitor global expression profile changes of Candida albicans genes in response to terbinafine treatment, and we anticipated a panoramic view of the responses of Candida albicans cells to the representatives of allylamine antifungal agents at the molecular level in an effort to identify drug class-specific and mechanism-independent changes in gene expression. METHODS: Candida albicans strain ATCC 90028 was exposed to either medium alone or terbinafine at a concentration equivalent to the 1/2 minimal inhibitory concentrations (MICs, 4 mg/L) for 90 minutes. RNA was isolated and gene expression profiles were compared to identify the changes in the gene expression profile using a cDNA microarray analysis. Differential expression of 10 select genes detected by cDNA microarray analysis was confirmed by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: A total of 222 genes were found to be responsive to terbinafine, including 121 up-regulated genes and 101 down-regulated genes. These included genes encoding membrane transport proteins belonging to the members of the ATP-binding cassette (ABC) or major facilitator superfamily (MFS; CDR1, AGP2, GAP6, PHO84, HOL3, FCY23, VCX1), genes involved in stress response and detoxification (CDR1, AGP2, HOL3), and gene involved in the ergosterol biosynthesis pathway (ERG12). The results of semi-quantitative RT-PCR were consistent with that of the cDNA microarray analysis. CONCLUSIONS: The up-regulation of the gene encoding the multidrug resistance efflux pump CDR1 may contribute to the terbinafine resistance in Candida albicans. However, the precise roles of other affected genes remain unclear, further studies of these genes and their respective products that play roles in the context of antifungal resistance are warranted.

Microwave endometrial ablation for menorrhagia in patients with systemic disorders.

OBJECTIVE: To evaluate the effectiveness of microwave endometrial ablation (MEA) in the treatment of menorrhagia in patients with severe systemic disease or medical conditions. METHODS: Forty-two menorrhagic women undergoing systemic disorders with failure of medical management were treated with MEA under local or general anesthesia, and were followed-up for 1 year. RESULTS: The women had a mean age of 39.4 years (range, 17-49). The procedure was successfully completed in all patients, and no intraoperative complications occurred. Two cases died of their primary severe medical diseases within 2 months of treatment but these cases were not associated with MEA. Among the remaining 40 patients, 24 (60.0%) had amenorrhea within 12 months. The duration of hospitalization and the amount of blood transfusion were significantly reduced after treatment, and the quality of life of these patients was improved significantly. CONCLUSIONS: MEA is a safe and effective treatment for the management of severe menorrhagia in patients undergoing systemic illness or severe medical conditions.

[Effects of Klebsiella pneumoniae on the expression of cytokines by polymorphonuclear neutrophil granulocytes in mice].

OBJECTIVE: To observe the effect of Klebsiella pneumoniae (Kle.p) on the expression of cytokines in mouse polymorphonuclear neutrophil granulocytes (PMNs). METHODS: Mouse models of acute pneumonia were induced by intranasal inoculation of 30 microl PBS containing 1 x 10(7) Kle.p (heat-inactivated at 60 degrees C for 1.5 h). The mRNA and protein expressions of the cytokines in the isolated and purified PMNs were assayed by reverse transcriptional (RT)-PCR, Western blotting and enzyme- linked immunosorbent assay (ELISA), respectively. RESULTS: Kle.p infection induced the mRNA expressions of tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta in the early stage, and TNF-alpha mRNA reached its peak at 6 h and IL-1beta mRNA at 12 h after infection, both of which decreased 24 h after infection. Western blot analysis showed that the protein expressions of TNF-alpha and IL-1beta were the highest at 6 h, and decreased obviously at 24 h after infection. ELISA showed that the secretion of TNF-alpha peaked at 6 h and decreased obviously at 24 h after infection, while the peak of IL-1beta production occurred at 12 h and maintained the high level till 24 h after the infection. CONCLUSION: TNF-alpha may participate in the immune defense of PMN against Kle.p infection mainly in the early stage of infection while IL-1beta mainly in the later stage.

[Study on the relationship between genesis and development of cervical cancer and the infection of human papillomavirus type 16/18, human herpesvirus II and cytomegalovirus].

OBJECTIVE: To investigate the correlation between genesis and the development of cervical cancer and infection of human papillomavirus (HPV) type 16/18, human herpesvirus II (HSV- II) and cytomegalovirus(CMV). METHODS: Different viruses were determined by polymerase chain reaction in 156 specimens of uterine including cervix 43 cervical cancer specimens,47 cervical intraepithelial neoplasia (CIN) specimens, 56 cervicitis specimens and 10 normal cervix specimens. RESULTS: (1) Positive rates on different viruses: the positive rates of HSV- II, HPV16/18 and CMV were declining in the cervical cancer specimens, CIN specimens or CIN III specimens and CIN I - II specimens, with significant differences. (2)Positive rate and grading, staging and histogenesis of cervical cancer on different viruses as well as positive rates of HPV16/18 in II staging cervical cancer specimens were significantly higher than that in I staging cervical cancer specimens while positive rates of HPV16/18 and HSV- II in high differentiation of cervical cancer specimens were significantly higher than those with medium differentiation from cervical cancer specimens. Positive rates of CMV did not seem to correlate with positive rate of HSV- II and CMV was not correlated to grading, staging or histogenesis of cervical cancer. (3)Copies of infected virus, HSV-II and HPV16/18 showing cervical cancer>CIN> cervicitis while with CMV:cervical cancer>CIN. (4) There were mixed infections of different viruses as HPV16/18 + HSV- II > HPV16/18 + CMV seen in the study. CONCLUSION: HPV 16/18, HSV- II and CMV infection were closely related to the genesis of cervical cancer and quantity of viruses which might have played an important role in carcinogenesis of cervical lesions.

Immunomodulating effects of cefodizime on Klebsiella pneumoniae-stimulated neutrophils.

To explore the immunoregulating effects of cefodizime on neutrophils and its mechanisms, we detected the expression of some cytokines secreted by neutrophils after heat-killed Klebsiella pneumoniae induced inflammation. We analyzed the changes of signal transduction in this process by detecting the mRNA expression of toll like receptor 4 (TLR4) and the inhibitor factor of kappaBalpha (I-kappaBalpha) expressed by neutrophils. The activity of nuclear factor kappaB (NF-kappaB) in neutrophils was also assayed by electrophoretic mobility shift assay (EMSA). We found cefodizime increased neutrophil production of TNF-alpha, IL-1beta in the early stage of inflammation, which was in agreement with the enhanced mRNA expression of TLR4 and DNA-binding activity of NF-kappaB. Taken together, the immunomodulating effects of cefodizime on cytokine production of K. pneumoniae stimulated neutrophils is possibly due to its regulation of TLR4 mRNA expression and DNA binding activities of NF-kappaB through the LPS-TLR4-NF-kappaB pathway.

[Investigation on the antibiotic resistance and risky factors of nosocomial infections caused by Stenotrophomonas maltophilia].

OBJECTIVE: To investigate the antibiotic resistance and risky factors of nosocomial infections caused by Stenotrophomonas maltophilia, so as to help elucidate the difference of drug resistance between metallic beta-lactamase (MBL) producing and non-MBL producing strains. METHODS: Standard agar dilution method of NCCLS was employed in the isolation of 36 strains of Stenotrophomonas maltophilia from patients with nosocomial infection with respect to their in vitro antibiotic resistance to 18 kinds of antibiotics. MBL strains were identified by MBL-E test method. RESULTS: Stenotrophomonas maltophilia in our hospital was mainly identified in the lower respiratory tract (88.9%), in which 88.2% (30/34) of the patients had serious original diseases, 50% of whom had received Imipenem/cilastatin sodium treatment. Thirty-six strains of Stenotrophomonas maltophilia were susceptible to new types of fluoquinolone antibiotics, i.e. Sparfloxacin, levofloxacin, gatifloxacin and doxycycline, with inhibitory rate ranging 97.2%, 94.4%, 91.7% to 83.3%, respectively. They could also be inhibited by SMZ/TMP and Ticarcillin/Lavulanic acid with inhibitory rate of 63.9% and 58.3%, respectively. There were 16 strains out of 36 of MBL bacteria with complete resistance to Imipenem/cilastatin sodium, but with higher susceptibility to aztreonam than those non-MBL producing strains. CONCLUSION: The nosocomial infection in our hospital caused by Stenotrophomonas maltophilia seemed to be related with severe primary disease and the use of Imipenem/cilastatin sodium. The newly developed fluoroquinolones possessed powerful antibacterial potency on Stenotrophomonas maltophilia found in nosocomial infection.

[Cloning and expression of metallo beta-lactamase encoding gene found in a clinical isolate of Stenotrophomonas maltophilia].

OBJECTIVE: To learn Molecular characterization of metallo beta-lactamase (MBL) found in a clinical isolate of Stenotrophomonas maltophilia and confirm the role of MBL played in the antimicrobial-resistance of S. maltophilia by sequencing the encoding genes of the metallo beta-lactamase and construct the prokaryotic expression vector carrying the MBL gene and expressed in E. coli BL21. METHODS: The blaMBL gene was amplified by PCR and cloned into pMD18-T plasmid. The recombination was subcloned into pET32a+ plasmid and expressed in E. coli BL21. The susceptibility between expression vectors and strain 750 to antibiotics were compared. RESULTS: The 867 bp DNA fragment of MBL encoding gene was amplified from the strain 750 by PCR and sequenced. The gene was 99.31% homologous to blaS and blaL1 of MBL L1. After being transformed into the E. coli BL21 and induced with lmM IPTG, a recombinant protein of about 48 kDa was expressed in the pET32a+-blaMBL system. The susceptibility of pET32a+-blaMBL system and strain 750 showed MIC 12 mg/L and 128 mg/L to imipenem and MIC 2 mg/L and 2 mg/L to ceftazidime, respectively. CONCLUSION: The MBL produced by strain 750 was similar to the that in strain ULA511. The difference of MIC to imipenem between wild strain and E. coli BL21 transformant indicated that other unclear mechanism involving in imipenem resistance in the strain.