Computational prediction of crucial genes involved in gonorrhea infection and neoplastic cell transformation: A multiomics approach.

Neisseria gonorrheae, the causative agent of genitourinary infections, has been associated with asymptomatic or recurrent infections and has the potential to form biofilms and induce inflammation and cell transformation. Herein, we aimed to use computational analysis to predict novel associations between chronic inflammation caused by gonorrhea infection and neoplastic transformation. Prioritization and gene enrichment strategies based on virulence and resistance genes utilizing essential genes from the DEG and PANTHER databases, respectively, were performed. Using the STRING database, protein‒protein interaction networks were constructed with 55 nodes of bacterial proteins and 72 nodes of proteins involved in the host immune response. MCODE and cytoHubba were used to identify 12 bacterial hub proteins (murA, murB, murC, murD, murE, purN, purL, thyA, uvrB, kdsB, lpxC, and ftsH) and 19 human hub proteins, of which TNF, STAT3 and AKT1 had high significance. The PPI networks are based on the connectivity degree (K), betweenness centrality (BC), and closeness centrality (CC) values. Hub genes are vital for cell survival and growth, and their significance as potential drug targets is discussed. This computational study provides a comprehensive understanding of inflammation and carcinogenesis pathways that are activated during gonorrhea infection.

The nitrone spin trap 5,5-dimethyl-1-pyrroline N-oxide dampens lipopolysaccharide-induced transcriptomic changes in macrophages.

OBJECTIVE: M1-like inflammatory phenotype of macrophages plays a critical role in tissue damage in chronic inflammatory diseases. Previously, we found that the nitrone spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO) dampens lipopolysaccharide (LPS)-triggered inflammatory priming of RAW 264.7 cells. Herein, we tested whether DMPO by itself can induce changes in macrophage transcriptome, and that these effects may prevent LPS-induced activation of macrophages. MATERIALS AND METHODS: To test our hypothesis, we performed a transcriptomic and bioinformatics analysis in RAW 264.7 cells incubated with or without LPS, in the presence or in the absence of DMPO. RESULTS: Functional data analysis showed 79 differentially expressed genes (DEGs) when comparing DMPO vs Control. We used DAVID databases for identifying enriched gene ontology terms and Ingenuity Pathway Analysis for functional analysis. Our data showed that DMPO vs Control comparison of DEGs is related to downregulation immune-system processes among others. Functional analysis indicated that interferon-response factor 7 and toll-like receptor were related (predicted inhibitions) to the observed transcriptomic effects of DMPO. Functional data analyses of the DMPO + LPS vs LPS DEGs were consistent with DMPO-dampening LPS-induced inflammatory transcriptomic profile in RAW 264.7. These changes were confirmed using Nanostring technology. CONCLUSIONS: Taking together our data, surprisingly, indicate that DMPO by itself affects gene expression related to regulation of immune system and that DMPO dampens LPS-triggered MyD88- and TRIF-dependent signaling pathways. Our research provides critical data for further studies on the possible use of DMPO as a structural platform for the design of novel mechanism-based anti-inflammatory drugs.

ß-keto amyrin isolated from Cryptostegia grandiflora R. br. inhibits inflammation caused by Daboia russellii viper venom: Direct binding of ß-keto amyrin to phospholipase A2.

The search for mechanism-based anti-inflammatory therapies is of fundamental importance to avoid undesired off-target effects. Phospholipase A2 (PLA2) activity is a potential molecular target for anti-inflammatory drugs because it fuels arachidonic acid needed to synthesize inflammation mediators, such as prostaglandins. Herein, we aim to investigate the molecular mechanism by which ß-keto amyrin isolated from a methanolic extract of Cryptostegia grandiflora R. Br. Leaves can inhibit inflammation caused by Daboia russellii viper (DR) venom that mainly contains PLA2. We found that ß-keto amyrin neutralizes DR venom-induced paw-edema in a mouse model. Molecular docking of PLA2 with ß-keto amyrin complex resulted in a higher binding energy score of -8.86 kcal/mol and an inhibition constant of 611.7 nM. Diclofenac had a binding energy of -7.04 kcal/mol and an IC50 value of 620 nM, which predicts a poorer binding interaction than ß-keto amyrin. The higher conformational stability of ß-keto amyrin interaction compared to diclofenac is confirmed by molecular dynamics simulation. ß-keto amyrin isolated from C. grandiflora inhibits the PLA2 activity contained in Daboia russellii viper venom. The anti-inflammatory property of ß-keto amyrin is due to its direct binding into the active site of PLA2, thus inhibiting its enzyme activity.

Exotoxins secreted by Clostridium septicum induce macrophage death: Implications for bacterial immune evasion mechanisms at infection sites.

The induction of macrophage death is considered a potential mechanism by which components secreted by Clostridium septicum are used to evade the innate immune response and cause tissue damage. This study aimed to determine the effects of partially purified fractions of extracellular proteins secreted by C. septicum on the death of mouse peritoneal macrophages. Elicited mouse peritoneal macrophages were incubated with partially purified fractions of proteins secreted by C. septicum into the culture medium. After incubation, the protein fraction with a molecular weight ≥100 kDa caused significant cell death in macrophages, altered cell morphology, increased the expression of markers of apoptosis and autophagy, and increased the expression (protein and mRNA) of IL-10 and TNFα. Our data suggest that the proteins secreted by C. septicum (MW, ≥100 kDa) induce cell death in macrophages by promoting autophagy-triggered apoptosis. This study may contribute to our understanding of the molecular mechanism of immune evasion by C. septicum at the infection site.

Marmesin and Marmelosin Interact with the Heparan Sulfatase-2 Active Site: Potential Mechanism for Phytochemicals from Bael Fruit Extract as Antitumor Therapeutics.

Human heparan sulfatase-2 (HSULF-2) is an oncoprotein overexpressed in the surface of all types of tumor cells and its activity plays a critical role in cancer survival and progression. Our previous studies have shown that bael fruit extract, containing marmesin and marmelosin, inhibits the HSULF-2 activity and kills breast tumor cells, but the mechanism of these processes remains fairly known mainly because the HSULF-2's 3D structure is partially known. Herein, we aimed at providing an in silico molecular mechanism of the inhibition of human HSULF-2 by phytochemicals from bael fruit extract. Pharmacokinetic parameters of the main phytochemicals contained in the bael fruit extract, sequence-based 3D structure of human HSULF-2, and the interaction of bael fruit's phytochemicals with the enzyme active site was modeled, evaluated, and verified. Docking studies revealed marmesin and marmelosin as potential inhibitors with binding score -8.5 and -7.7 Kcal/mol; these results were validated using molecular dynamics simulations, which exhibited higher stability of the protein-ligand complexes. Taking together, with our earlier in vitro data, our computational analyses suggest that marmesin and marmelosin interact at the active site of HSULF-2 providing a potential mechanism for its inhibition and consequent antitumor activity by phytochemicals contained in the bael fruit extract.

Anti-oxidant and anti-inflammatory effect of polar extracts obtained from waste product of wine making.

A lot of diseases are characterized by an increased inflammatory response with an exacerbated production of free radicals. The anti-inflammatory effect of different compounds with antioxidant capacity, as polyphenols present in grape is well known. Therefore, the objective is to evaluate the anti-oxidant and anti-inflammatory activity of waste product of wine production.Six different non-toxic-marc-polar extracts from Malbec and Syrah grape varieties were obtained, their total phenol and flavonoid content were evaluated, and their antioxidant and anti-inflammatory activity were determined.High content of total phenols and flavonoids were found mainly in extracts obtained from Syrah (80.51 ± 16.63 g equivalent to gallic acid/100 g and 25.47 ± 3.33 g equivalent to quercetin/100 g). In addition, they had a high antioxidant effect (above 88.5% of ABTS inhibition by Syrah extracts). Finally, all extracts decreased the nitric oxide (NO) production, but this was more accented when extract from Syrah obtained by infusion was used, which decreased NO levels to baseline (4.46 µM).Taking together, our results show the potential pharmaceutical use of waste product of wine making to prevent or to treat diseases which inflammatory response is exacerbated.

A chronic high-fat diet causes sperm head alterations in C57BL/6J mice.

A chronic-positive energetic balance has been directly correlated with infertility in men, but the involved mechanisms remain unknown. Herein we investigated weather in a mouse model a chronic feeding with a diet supplemented with chicken fat affects sperm head morphology. To accomplish this, we fed mice for 16 weeks with either control food (low-fat diet, LFD) or control food supplemented with 22% chicken fat (high-fat diet, HFD). At the end of the feeding regimen, we measured: redox and inflammatory changes, cholesterol accumulation in testis and analyzed testicular morphological structure and ultra-structure and liver morphology. We found that the mice fed HFD resembled some features of the human metabolic syndrome, including systemic oxidative stress and inflammation, this group showed an increment in the following parameters; central adiposity (adiposity index: 1.07 ± 0.10 vs 2.26 ± 0.17), dyslipidemia (total cholesterol: 153.3 ± 2.6 vs 175.1 ± 8.08 mg/dL), insulin resistance (indirect Insulin resistance index, TG/HDL-c: 2.94 ± 0.33 vs 3.68 ± 0.15) and fatty liver. Increased cholesterol content measured by filipin was found in the testicles from HFD (fluorescence intensity increase to 50%), as well as an alteration of spermiogenesis. Most remarkably, a disorganized manchette-perinuclear ring complex and an altered morphology of the sperm head were observed in the spermatozoa of HFD-fed mice. These results add new information to our understanding about the mechanisms by which systemic oxidative stress and inflammation may influence sperm-head morphology and indirectly male fertility.

Therapeutic targets to reduce the contribution of pulmonary neutrophilic inflammation towards obesity-associated co-morbidities: a mini-review.

Epidemiology and experimental models have shown a close link between adipose tissue inflammation, systemic inflammation and pulmonary neutrophilic inflammation, which predispose obese patients to pulmonary diseases, obesity-associated co-morbidities and cancer. Increased content and activation of neutrophils in the lung microvasculature, resulting from peripheral activation of neutrophils, and increased adhesion of neutrophils to the lung microvasculature are important factors explaining the increased susceptibility of obese patients towards respiratory diseases and loss of insulin sensitivity. Mechanism-based therapies to break this link are urgently needed to reduce pulmonary damage in obesity, due to the growing prevalence of obesity world-wide. Current research suggests that these approaches should be focused on, one or more of the following: reduction of macrophage activation at the adipose tissue, healthy growing of adipose tissue by induction of Nrf-2, inhibition of NF-κB activation, reduction of circulating neutrophil activation, blocking adhesins/selectins, inhibition of neutrophil activation by targeting NADPH oxidase-2 activation, inhibition of myeloperoxidase activity and scavenging of hypochlorous acid. These strategies are expected to reduce adipose tissue inflammation, peripheral inflammation, pulmonary neutrophilic inflammation and obesity-associated co-morbidities.

Varied protocols of cadmium exposure produce different effects on nitric oxide production in macrophages.

Different protocols of cadmium (Cd) exposure in non-cytotoxic conditions (i.e. 10 microM Cd for 18 h), and their effect on nitric oxide (NO) generation induced by NO inductor agents (NOIA) in peritoneal macrophages (pM) were studied. In all cases, NOIA (i.e. bacterial lipopolysaccharide [LPS], phorbol ester [PMA], okadaic acid [OA] or their combinations [LPS/OA] and [LPS/PMA]) were added at the beginning of the first incubation, only. Simultaneously exposure with 10 microM Cd enhanced NO generation and inducible NO synthase (iNOS) expression evoked by LPS, OA, PMA; those induced by LPS/PMA were not modified; and those caused by LPS/OA in relation to culture without Cd (medium) decreased. Double incubation, either with or without Cd (Cd+Cd or medium+medium), or Cd added at the start of the first or second incubation only (Cd+medium or medium+Cd), were tested. After the second incubation, medium+Cd protocol produced the highest NO generation in relation to other exposure protocols. When NO production was measured at the end of the second incubation, Cd+medium protocol enhanced NO production induced by OA, and LPS/OA, while medium+Cd protocol enhanced the response to LPS, PMA, and LPS/OA, in both cases in relation to the first incubation. Cd+Cd incubation protocol decreases the response to all NOIA in relation to another protocols. Cd effect on NO generation in macrophages is dependent on protocol and timing of exposure.

Biphasic effect of cadmium in non-cytotoxic conditions on the secretion of nitric oxide from peritoneal macrophages.

The effects of cadmium (Cd) in non-cytotoxic conditions on the nitric oxide (NO) production in peritoneal macrophages (pM) were studied. Peritoneal macrophages from Balb/c mice were incubated over 18 h with 5, 10, 20, or 25 microM Cd2+ (as CdCl2 21:2 H2O) in the culture medium. Concentrations of 20 microM Cd2+ and over had cytotoxic effects, measured by MTT assay. Cell viability with 10 microM Cd2+ in the medium was above 90% after 18 h of incubation, and above 80% after 72 h. At this same Cd2+ concentration, NO production increased from 6 to 18 h. At 24 h production decreased but was still above control levels. At 48 h production NO was near control levels, and continued to decrease until the end of the experiment (72 h). NO levels produced with Cd2+ concentrations of 5, 10 and 20 microM in the medium were above the control at 18 h. NO production and lipoperoxidation increased simultaneously after 18 h with 10 microM of Cd in the medium. Amounts of inducible nitric oxide synthase (iNOS) protein and iNOS activity also increased. At a concentration of 10 microM Cd has a biphasic effect on NO production over time.

Modulation of arachidonic acid turnover in macrophages by cadmium.

The effects of cadmium (Cd) induced redox changes on arachidonic acid (AA) turnover in mouse resident peritoneal macrophages (pM) were studied. The pre-incubation of pM in a medium containing glutathione (GSH, 0.1 or 1 mM) for 6 h protects pM from loss of viability and AA uptake diminution induced by Cd with regard to non pre-incubated cultures. The exposure of macrophages to Cd 10 microM decreases AA uptake within 2 h and increases AA release in relation to non-exposed macrophages. It also enhances AA mobilization and reactive oxygen species (ROS) release induced by okadaic acid and opsonized zimosan and decreases those induced by lipopolysaccharide, but does not modify either AA mobilization or ROS release induced by phorbol ester. These results might suggest that redox changes induced by Cd produce an important impact on AA turnover in macrophages; information that is relevant in the understanding of the cellular toxicity of this metal.

Gastrostomía endoscópica percutánea: experiencia de 5 años / Percutaneous endoscopic gastrostomy: 5 years experience

309 gastrostomías percutáneas sucesivas fueron realizadas en 206 pacientes por un solo operador en el Servicio de Endoscopía del Hospital DIPRECA durante los años 1993 al 1998. La edad promedio de los pacientes fue de 71 años (18-91 años). El 15 por ciento requirió al menos de un recambio. El seguimiento fluctuó entre 4 y 20 meses. Hubo un 3.87 por ciento de morbilidad y 0 por ciento de mortalidad. La indicación más frecuente son las enfermedades neurológicas