Colonization of multidrug-resistant Gram-negative bacteria increases risk of surgical site infection after hemorrhoidectomy: a cross-sectional study of two centers in southern China.

PURPOSE: The present study aims to determine the rectoanal colonization rate and risk factors for the colonization of present multidrug-resistant bacteria (MDRBs). In addition, the relationship between MDRB colonization and surgical site infection (SSI) following hemorrhoidectomy was explored. METHODS: A cross-sectional study was conducted in the Department of Colorectal Surgery of two hospitals. Patients with hemorrhoid disease, who underwent hemorrhoidectomy, were included. The pre-surgical screening of multidrug-resistant Gram-negative bacteria (MDR-GNB) colonization was performed using rectal swabs on the day of admission. Then, the MDRB colonization rate was determined through the rectal swab. Logistic regression models were established to determine the risk factors for MDRB colonization and SSI after hemorrhoidectomy. A p-value of < 0.05 was considered statistically significant. RESULTS: A total of 432 patients met the inclusion criteria, and the MDRB colonization prevalence was 21.06% (91/432). The independent risk factors for MDRB colonization were as follows: patients who received ≥ 2 categories of antibiotic treatment within 3 months (odds ratio (OR): 3.714, 95% confidence interval (CI): 1.436-9.605, p = 0.007), patients with inflammatory bowel disease (IBD; OR: 6.746, 95% CI: 2.361-19.608, p < 0.001), and patients with high serum uric acid (OR: 1.006, 95% CI: 1.001-1.010, p = 0.017). Furthermore, 41.57% (37/89) of MDRB carriers and 1.81% (6/332) of non-carriers developed SSIs, with a total incidence of 10.21% (43/421). Based on the multivariable model, the rectoanal colonization of MDRBs (OR: 32.087, 95% CI: 12.052-85.424, p < 0.001) and hemoglobin < 100 g/L (OR: 4.130, 95% CI: 1.556-10.960, p = 0.004) were independently associated with SSI after hemorrhoidectomy. CONCLUSION: The rectoanal colonization rate of MDRBs in hemorrhoid patients is high, and this was identified as an independent risk factor for SSI after hemorrhoidectomy.

The study of neuroprotective effect of ferulic acid based on cell metabolomics.

Ferulic acid (FA), a naturally derived phenolic compound, has antioxidant and antidepressant-like effects. It is still a challenge to study its mechanism due to the complexity of the pathophysiology of depression. In this study, ultra-high performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS) was used to perform metabolomics studies based on biochemical changes in differentiated rat pheochromocytoma (PC12) cells treated with corticosterone-induced neurological damage after FA treatment. A total of 31 metabolites were identified as potential biomarkers for corticosterone-induced PC12 cells injury. Among them, 24 metabolites were regulated after FA treatment. Pathway analysis revealed that these metabolites were mainly involved in the amino acid metabolism, energy metabolism and glycerophospholipid metabolism. In addition, based on the results of metabolomics, three cell signaling pathways related to glutamate were discovered. To further study the interactions between FA and major targets in three signaling pathways, a molecular docking method was employed. The results showed that FA had the strongest binding power with protein kinase B (AKT). Furthermore, the result of mRNA changes analyzed by quantitative real time RT-PCR indicated that AKT and protein kinase A (PKA) in the signaling pathway were up regulated after treatment with FA compared with model group. This study shows that strategies based on cell metabolomics associated with molecular docking and molecular biology is a helpful tool to elucidate the neuroprotective mechanism of FA.

Investigation of the hepatoprotective effect of Corydalis saxicola Bunting on carbon tetrachloride-induced liver fibrosis in rats by 1H-NMR-based metabonomics and network pharmacology approaches.

Liver fibrosis is a common consequence of chronic liver diseases resulting from multiple etiologies. Furthermore, prolonged unresolved liver fibrosis may gradually progress to cirrhosis, and eventually evolve into hepatocellular carcinoma (HCC). Corydalis saxicola Bunting (CS), a type of traditional Chinese folk medicine, has been reported to have hepatoprotective effects on the liver. However, the exact mechanism of how it cures liver fibrosis requires further elucidation. In this work, an integrated approach combining proton nuclear magnetic resonance (1H-NMR)-based metabonomics and network pharmacology was adopted to elucidate the anti-fibrosis mechanism of CS. Metabonomic study of serum biochemical changes by carbon tetrachloride (CCl4)-induced liver fibrosis in rats after CS treatment were performed using 1H-NMR analysis. Metabolic profiling by means of partial least squares-discriminate analysis (PLS-DA) indicated that the metabolic perturbation caused by CCl4 was reduced after CS treatment. As a result, lipids, leucine, alanine, acetate, O-acetyl-glycoprotein and creatine were significantly restored after CS treatment, which regulated valine, leucine and isoleucine metabolism; arginine and proline metabolism; lipid metabolism and pyruvate metabolism. Additionally, 157 potential targets of CS and 265 targets of liver fibrosis were identified by means of network pharmacology. Subsequently, 5 target proteins, which are the intersection of potential CS targets and liver fibrosis targets, indicated that CS has potential anti-fibrosis effects through regulating alanine aminotransferase (ALT) activity, the farnesoid X receptor (FXR), cyclooxygenase-2 (COX-2), matrix metalloproteinase-1 (MMP-1) and angiotensinogen. Chelerythrine and sanguinarine were the potential active compounds in CS for treating liver fibrosis through regulating ALT activity. This study is the first report to study the anti-fibrosis effects of CS on the basis of combining a metabonomics and network pharmacology approaches, and it may be a potentially powerful tool to study the efficacy and mechanisms of traditional Chinese folk medicines.

Rap2a serves as a potential prognostic indicator of renal cell carcinoma and promotes its migration and invasion through up-regulating p-Akt.

Rap2a, a member of the small GTPase superfamily, belongs to Ras superfamily, and its function in cancer progression is still poorly understood. Our previous study indicated that the ectopic expression of Rap2a enhanced the migration and invasion ability of lung cancer cells. However, its expression and molecular mechanism on renal cell carcinoma (RCC) have not been characterized. This study explored the clinical significance and biological function of Rap2a in human RCC. The clinical relevance of Rap2a in RCC was evaluated by immunohistochemical staining using tissue microarray. Our data showed that Rap2a expression was dramatically increased in RCC tissues compared with normal renal tissues. The ectopic expression of Rap2a enhanced the migration and invasive ability of cancer cells. In contrast, downregulation of Rap2a inhibited cell invasion. Rap2a had no effect on the proliferation of RCC cell lines. Meanwhile, Rap2a can regulate the phosphorylation level of Akt in vitro. In vivo studies also showed that Rap2a positively regulated metastasis of renal cancer cells and the expression of p-Akt. These findings indicate that Rap2a promotes RCC metastasis and may serve as a candidate RCC prognostic marker and a potential therapeutic target.

Rap2a is a novel target gene of p53 and regulates cancer cell migration and invasion.

The p53 transcription factor is a critical regulator of the cell cycle, DNA repair, and apoptosis. Recent evidences suggest that p53 may contribute to the regulation of cell invasion and migration. Rap2a, a member of the small GTPase superfamily, mediates diverse cellular events such as cell adhesion, migration and proliferation through various signaling pathways. In this study, we identify that Rap2a is a novel target of p53 and is induced upon DNA damage in a p53-dependent manner. Upon DNA damage, p53 directly binds to the promoter of Rap2a and activates its transcription. We show that Rap2a is significantly upregulated in many types of tumors. In addition, the ectopic expression of Rap2a enhances the migration and invasive ability of cancer cells and increases activities of matrix metalloproteinase MMP2 and MMP9. In contrast, the inactivation of Rap2a inhibits cell invasion and activities of MMP2 and MMP9. We also show that Rap2a regulates the phosphorylation level of Akt. Collectively, our results show that ectopic expression of Rap2a has a key role in enhancing migration, invasion and metastasis by upregulating p-Akt.

p21-Activated kinase 5 affects cisplatin-induced apoptosis and proliferation in hepatocellular carcinoma cells.

p21-Activated kinase 5 (PAK5) is the last identified member of the PAK family. The PAKs are highly conserved serine/threonine and effector proteins for Cdc42 and Rac and are essential in regulating cell motility and survival. Previous studies have demonstrated that PAK5 played a pivotal role in apoptosis, proliferation, cancer migration, and invasion. However, the biological function of PAK5 in hepatocellular carcinoma, as well as its underlying mechanism, still remains to be fully elucidated. In the present study, we demonstrated that PAK5 markedly inhibited cisplatin-induced apoptosis and promoted cell proliferation in hepatocellular carcinoma cells. Moreover, our results showed that overexpression of PAK5 contributed to cell cycle regulation. In order to elucidate the underlying mechanism of PAK5 on cisplatin-induced apoptosis and cell cycle regulation, we also examined the protein expressions of chk2 and p-chk2. In summary, our study investigated the role of PAK5 in cisplatin-induced cellular processes and provided evidence of its underlying mechanism.

Gamma-aminobutyric acid induces tumor cells apoptosis via GABABR1·ß-arrestins·JNKs signaling module.

Gamma-aminobutyric acid (GABA), an inhibitory neurotransmitter in central nervous system, has yet been found to widely exist in tumor tissues to regulate tumor cells growth. However, the function of GABA on inducing tumor cells apoptosis and the potential mechanism are still unclear. In order to detect whether GABA via GABAB receptor GABABR1 would activate c-Jun N-terminal kinases (JNKs) to promote tumor cells apoptosis, co-immunoprecipitation assay was used to investigate the association of ß-arrestins with GABABR1 and JNKs in the different four cancer cell lines. Our observation demonstrated that ß-arrestins, in addition to their role in G protein-coupled receptors desensitization, had an additional function as adapter proteins to recruit JNKs to GABABR1, thereby conferring distinct enzymatic activities upon the receptor, which may trigger JNKs signal pathway involved in the regulation of cellular growth. Activated JNKs subsequently phosphorylated downstream c-Jun to transcribe a wide variety of pro-apoptotic genes. Additionally, GABA up-regulated the ratio of pro-apoptotic protein Bax to anti-apoptotic protein Bcl-2, and thus facilitated caspase-3 cleavage, leading to tumor cells apoptosis in a mitochondrial-dependent pathway. In contrast, GABABR antagonist CGP35348 reversed GABA-induced JNKs phosphorylation and its downstream proteins activation, which consequently restrained tumor cells apoptosis. Taken together, our study suggested that GABA via its receptor GABABR1 recruited ß-arrestins to facilitate the activation of JNKs cascade, resulting in tumor cells growth inhibition.

Ubiquitination of p53 is involved in troglitazone induced apoptosis in cervical cancer cells.

Peroxisome proliferator-activated receptor gamma (PPAR-γ), a ligand-dependent nuclear transcription factor, has been found to widely exist in tumor tissues and plays an important role in affecting tumor cell growth. In this study, we investigated the effect of PPAR-γ on aspects of the cervical cancer malignant phenotype, such as cell proliferation and apoptosis. Cell growth assay, Western blotting, Annexin V and flow cytometry analysis consistently showed that treatment with troglitazone (TGZ, a PPAR-γ agonist) led to dose-dependent inhibition of cervical cancer cell growth through apoptosis, whereas T0070907 (another PPAR-γ antagonist???) had no effect on Hela cell proliferation and apoptosis. Furthermore, we also detected the protein expression of p53, p21 and Mdm2 to explain the underlying mechanism of PPAR-γ on cellular apoptosis. Our work, finally, demonstrated the existence of the TGZ-PPAR-γ-p53 signaling pathway to be a critical regulator of cell apoptosis. These results suggested that PPAR-γ may be a potential therapeutic target for cervical cancer.

ß-arrestin promotes c-Jun N-terminal kinase mediated apoptosis via a GABA(B)R·ß-arrestin·JNK signaling module.

Evidence is growing that the GABAB receptor, which belongs to the G protein-coupled receptor (GPCR) superfamily, is involved in tumorigenesis. Recent studies have shown that ß-arrestin can serve as a scaffold to recruit signaling protein c-Jun N-terminal knase (JNK) to GPCR. Here we investigated whether ß-arrestin recruits JNK to the GABAB receptor and facilitates its activation to affect the growth of cancer cells. Our results showed that ß-arrestin expression is decreased in breast cancer cells in comparison with controls. ß-arrestin could enhance interactions of the GABABR·ß-arrestin·JNK signaling module in MCF-7 and T-47D cells. Further studies revealed that increased expression of ß-arrestin enhances the phosphorylation of JNK and induces cancer cells apoptosis. Collectively, these results indicate that ß-arrestin promotes JNK mediated apoptosis via a GABABR·ß-arrestin·JNK signaling module.

Downregulation of PAK5 inhibits glioma cell migration and invasion potentially through the PAK5-Egr1-MMP2 signaling pathway.

PAK5 (p21 activated kinase 5) is upregulated in human colorectal carcinoma cells and is a known tumor promoter in carcinogenesis of the colon. Little is known regarding the mechanisms underlying the downstream targets of PAK5, and information concerning its biological significance in glioma is lacking. In this study, we investigated the effects of PAK5 on proliferation, migration, invasion, and apoptosis in human U87 and U251 glioma cells and examined the underlying molecular mechanism. We performed cell growth assays and cell cycle analysis to observe the cell proliferation. Flow cytometry analysis was performed to evaluate apoptosis, and in vitro scratch assays, cell migration assays, and gelatin zymography were performed to examine cell migration. Western blot analysis was performed to examine signal transduction in the cells. We demonstrated that suppression of PAK5 in glioma cells significantly inhibited cell migration and invasion. We also observed that suppression of PAK5 in human glioma cell lines inhibited cell growth because of G1 phase arrest. Additionally, flow cytometry and Western blot analysis indicated that PAK5 could inhibit cell apoptosis. These results suggest that the PAK5-Egr1-MMP2 signaling pathway is involved in tumor progression and may have a potential role in cancer prevention and treatment.

A sphingosine kinase-1 inhibitor, SKI-II, induces growth inhibition and apoptosis in human gastric cancer cells.

SKI-II has been reported as an inhibitor of sphingosine kinase 1 and has been extensively used to prove the involvement of sphingosine kinase and sphingosine-1-phosphate (Sphk1) in cellular processes. In the current study, we investigated the effects of SKI-II and its potential mechanisms in human gastric cancer SGC7901 cells. After treatment with SKI-II, cell growth, cell cycle distribution, apoptosis, expression of Sphk1, NF-κB, Bcl-2, Bax and p27 were assessed by MTT assay, flow cytometry, electron microscopy, immunocytochemistry and Western-blot assay, respectively. Our results showed that SKI-II markedly inhibited SGC7901 cell survival in a dose-dependent manner, reduced cell proliferation with accumulation of cells in the G0/G1 phase and induced apoptosis in the tumor cells. Furthermore, Western blotting and immunocytochemistry showed that the expression of p27 and Bax was increased significantly, but the expression of NF-κB, Bcl-2 and Sphk1 decreased by different degrees. These results indicate that SKI-II induced cell growth arrest and apoptosis. The increased apoptotic sensitivity of SGC7901 was correlated with NF-κB or Bcl-2/Bax activation.

PAK5-Egr1-MMP2 signaling controls the migration and invasion in breast cancer cell.

p21-activated kinases (PAKs) are activated by various extracellular stimuli and, in turn, activate other kinases by phosphorylating them at specific serine/threonine residues or through protein-protein interaction. As a recently identified member of the group B PAK family, the role of PAK5 in cancer is poorly understood. In this study, we investigated the effect of PAK5 on the malignant phenotype, such as proliferation, cell cycle, apoptosis, migration, and invasion. Cell growth assay and cell cycle analysis consistently showed that knockdown of PAK5 could significantly inhibit the proliferation of breast cancer cells. Wound healing assay. migration assay, and invasion assay showed that PAK5 promoted cell migration. Furthermore, in order to elucidate the underlying mechanism of PAK5 on cellular growth and migration, we examined the protein expressions of cyclin D1, p21, early growth response protein 1 (Egr1), and matrix metalloproteinase 2 (MMP2). Our work further reveals the PAK5-Egr1-MMP2 signaling pathway to be a critical regulator of cell migration and invasion. These results suggest that PAK5 may be a potential therapeutic target for breast cancer.

Differential expression of Bcl-2 and Bax during gastric ischemia-reperfusion of rats.

AIM: To investigate expression of Bcl-2 and Bax in gastric ischemia-reperfusion (GI-R) and involvement of extracellular signal-regulated kinase (ERK) 1/2 activation. METHODS: The GI-R model was established by ligature of the celiac artery for 30 min and reperfusion in Sprague-Dawley rats. Rats were assigned to groups in accordance with their evaluation period: control, 0, 0.5, 1, 3, 6, 24, 48, and 72 h. Expression and distribution of Bcl-2 and Bax proteins were analyzed by immunohistochemistry and western blotting in gastric tissue samples after sacrifice. RESULTS: Compared with controls, the percentage of positive cells and protein levels of Bcl-2 decreased in the early phases of reperfusion, reached its minimum at 1 h (P < 0.05); it then increased, reaching its peak at 24 h of reperfusion (P < 0.05). The pattern of Bax expression was opposite to that of Bcl-2. Bax expression increased after reperfusion, with its peak at 1 h of reperfusion (P < 0.05), and then it decreased gradually to a minimum at 24 h after reperfusion (P < 0.05). On the other hand, inhibition of activation of ERK1/2 induced by PD98059, a specific upstream MEK inhibitor, had significant effects on Bcl-2 and Bax in GI-R. Compared with GI-R treatment only at 3 h of reperfusion, PD98059 reduced the number of Bcl-2 positive cells (0.58% of R3h group, P < 0.05) and Bcl-2 protein level (74% of R3h group, P < 0.05) but increased the number of Bax-positive cells (1.33-fold vs R3h group, P < 0.05) and Bax protein level (1.35-fold of R3h group, P < 0.05). CONCLUSION: These results indicated that the Bcl-2 and Bax played a pivotal role in the gastric mucosal I-R injury and repair by activation of ERK1/2.

[Effects of genistein on contractility of isolated right ventricular muscles in guinea pig].

AIM: To study the effect of genistein (GEN) on contractility of isolated right ventricular muscles in guinea pig and its mechanisms. METHODS: Isolated guinea pig ventricular muscles were suspended in organ baths containing K-H solution.After an equilibration period, the effect of GEN on contraction of myocardium was observed. RESULTS: GEN and isoprenaline hydrochloride had the positive inotropic effects on contractity of myocardium. Meanwhile, the effect of GEN (1-100 micromol x L(-1)) was in dose-dependent manner. Propranolol (1 micromol x L(-1)) and verapamil hydrochloride (0.5 micromol x L(-1)) attenuated the positive inotropic effect of isoprenaline hydrochloride (1 micromol x L(-1)), but did not change the effect of GEN (50 micromol x L(-1)). Further more, the enhancement of the contraction induced by elevation of extracellular Ca2+ concentration in ventricular muscles had no change after pretreatment with GEN (1.10 micromol x L(-1)). In addition,the positive inotropic effect of GEN was inhibited partially by tamoxifen (1 micromol x L(-1)) and SQ22536 (1 micromol x L(-1)), also, could be attenuated by bpV (1 micromol x L(-1)). CONCLUSION: GEN has the positive inotropic effect on guinea pig ventricular muscles, which is not related to the activation of beta adrenoceptor, Ca2+ channel on cell membrane,but may involve in cAMP of intracellular signal transduction and tyrosine kinase pathway.

Effects of phytoestrogens and 17beta-estradiol on vasoconstriction elicited by reactive oxygen species.

To study the effects of different reactive oxygen species (ROS) on the resting tension of porcine coronary artery rings and to identify the effects of genistein (GEN), resveratrol (RES) and 17beta-estradiol (EST) on ROS-elicited vasoconstriction, porcine coronary rings were prepared and mounted in an organ bath and, after an equilibration period, the changes induced by the drugs were observed. Rings with intact endothelium showed an obvious but slow contraction after treatment with xanthine (100 microM)/xanthine oxidase (20 mU x mL(-1)) (X/XO) whereas endothelium-denuded rings showed no effects. H2O2 (200 microM) induced a fast and transient contraction in endothelium-denuded rings and failed to do so in intact-endothelium rings. Like superoxide dismutase (SOD, 200 U x mL(-1)), GEN (1 microM) and RES (1 microM) significantly inhibited contractile response evoked by X/XO, however in contrast to GEN and RES, EST (1 microM) had no obvious effect. GEN (30 microM) and RES (30 microM), like catalase (CAT, 800 U x mL(-1)), markedly attenuated the contraction elicited by H2O2. The results demonstrate that GEN and RES have distinct inhibitory effects on vasoconstriction induced by O2*- generated by X/XO and H2O2, and their actions are clearly greater than to that of EST.

Differential mechanisms involved in effects of genistein and 17-beta-estradiol on porcine coronary arteries.

The purpose of this work was to examine the differential mechanisms involved in relaxation induced by genistein and 17-beta-estradiol in isolated porcine coronary arteries. Similar to 17-beta-estradiol, genistein could dose-dependently relax 30 mM KCI-precontracted coronary artery rings. The pD2 values of genistein and 17-beta-estradiol were 4.91 +/- 0.13 and 4.98 +/- 0.12 respectively. Incubation with N-L-nitroarginine (L-NNA), endothelium removal or in the presence of a potent inhibitor of protein tyrosine phosphatase sodium orthovanadate did not affect the relaxation induced by genistein, but could partially reduce the vasorelaxation induced by 17-beta-estradiol. The relaxations induced by genistein and 17-beta-estradiol were unaffected by the estrogen receptor antagonist tamoxifen, the inhibitor of prostanoid synthesis indomethacin and the protein synthesis inhibitor, cycloheximide. In addition, both of genistein and 17-beta-estradiol could decrease the contractile responses of KCI, 5-HT and CaCl2, and shift their cumulative concentration-response curves rightward in a parallel manner. These findings suggest that the relaxant effects induced by genistein and 17-beta-estradiol are probably mainly due to inhibition of Ca2+ influx through voltage-dependent calcium channels (VDCCs), and are not related to sex hormone receptor and classical genomic activities. Also there is an interesting finding that the relaxing response of 17-beta-estradiol is partially endothelium-dependent, but that of genistein is not.

A study of mechanisms involved in vasodilatation induced by resveratrol in isolated porcine coronary artery.

The present study was designed to investigate the acute relaxing effect of phytoestrogen resveratrol on isolated porcine coronary arteries and to determine the mechanisms underlying its vasodilatation. Rings of porcine coronary arteries were suspended in organ baths containing Krebs-Henseleit solution, and then isometric tension was measured. Resveratrol concentration-dependently relaxed arterial rings precontracted with 30 mM KCl. The IC(50) value of resveratrol was 38.67+/-3.21 microM. Incubation with N(omega)-L-nitro-arginine (L-NNA), endothelium removal or the presence of a potent inhibitor of protein tyrosine phosphatase sodium orthovanadate partly decreased the relaxation induced by resveratrol. However, the relaxation induced by resveratrol was unaffected by the estrogen receptor antagonist tamoxifen, the inhibitor of prostanoid synthesis indomethacin, the antagonist of beta-adrenoceptors propranolol or the protein synthesis inhibitor, cycloheximide. In addition, resveratrol significantly decreased the contractile responses of 5-HT, KCl and CaCl(2), and shifted their cumulative concentration-response curves to the right. These results suggest that the mechanisms of vasorelaxation induced by resveratrol are heterogeneous, two mechanisms participating partially in the relaxation of porcine coronary artery were detected in the study, one being the nitric oxide released from the endothelium, the other causing inhibition of Ca(2+) influx, but estrogen receptors were not involved in resveratrol-induced relaxation.