[Pre-clinical in vivo evaluation study on a new Chinese-made surgical biopatch for atrial septum].

Objective: To evaluate the safety and effectiveness of a new Chinese-made surgical biopatch for atrial septum under the establishment of atrial septal defect animal model in miniature pigs. Methods: From June 2018 to April 2019, 26 pigs were divided into experimental group (15 pigs) and the control group (11 pigs). Animal models of atrial septal defect were established by traditional surgical methods. The to-be-evaluated and listed surgical biological patches (with a diameter of 10 mm) were implanted in the experimental group and the control group to repair the atrial septal defect. Cardiac ultrasound and blood examination of all animals were performed before and at 7, 30, 90, 180 days after operation, the results were analyzed with repetitive measurement and analysis of variance. At 90 days and 180 days after the operation, tissue samples were taken from animals after euthanasia. Pathological examination of heart and major organs were conducted. The independent sample t test and rank sum test were used to compare the data between the two groups, and the nonparametric was used to compare the patch calcification score between the two groups. Results: In total of 26 animals, 14 animals in the experimental group(6 at 90 days, 8 at 180 days) and 9 animals in the control group(4 at 90 days, 5 at 180 days) reached the end of the experiment. The other 3 animals (1 in the experimental group and 2 in the control group) died of arrhythmia, whole heart failure and right heart failure, the results of pathological examination showed that the causes of death were unrelated to the experimental materials. Cardiac ultrasound showed no patch leakage in all animals. There was no statistically significant difference in cardiac ultrasound and blood examination between the two groups at different time points after operation (all P>0.05). The pathological results showed that all the implants were intact and had good biocompatibility. There was no significant difference in the mean endothelialization rate between the experimental group and the control group at 90 and 180 days after operation ((80.8±29.1)% vs. (82.5±23.6)%, t=0.095, P=0.927; (78.8±36.4)% vs. (82.0±19.2)%, t=0.182, P=0.859) on 90 and 180 days, there was no significant difference in the patch calcification score between the two groups (1.00(1.25) vs. 2.00(0.75), Z=6.500, P=0.214; 0(0.75) vs. 1.00(2.00), Z=12.000, P=0.139). Conclusion: The new Chinese-made surgical biopatch for atrial septum has comparable safety and efficacy to that of the marketable patch in miniature pig atrial septal defect animal model.

Seven genes for the prognostic prediction in patients with glioma.

PURPOSE: Glioma is a common malignant tumor of the central nervous system, which is characterized by a low cure rate, high morbidity, and high recurrence rate. Consequently, it is imperative to explore some indicators for prognostic prediction in glioma. METHODS: We obtained glioma data from The Cancer Genome Atlas (TCGA). Differentially expressed genes (DEGs) were obtained by R software from TCGA data sets. Through Cox regression analysis, risk scores were obtained to assess the weighted gene-expression levels, which could predict the prognosis of patients with glioma. The validity and the prognostic value of this model in glioma were confirmed by the manifestation of receiver-operating characteristic (ROC) curves, area under the curve (AUC), and 5-year overall survival (OS). RESULTS: In total, 920 DEGs of transcriptome genes in glioma were extracted from the TCGA database. We identified a novel seven-gene signature associated with glioma. Among them, AL118505.1 and SMOC1 were positively related to the 5-year OS of patients with glioma, showing a better prognosis for glioma; however, RAB42, SHOX2, IGFBP2, HIST1H3G, and IGF2BP3 were negatively related to 5-year OS, displaying a worse prognosis. In addition, according to risk scores, AL118505.1 was also a protective factor, while others were risk factors. Furthermore, the expression levels of SHOX2, IGFBP2, and IGF2BP3 were significantly positively correlated with glioma grades. Receiver-operating characteristic (ROC) curve assessed the accuracy and sensitivity of the gene signature. Each of the seven genes for patients with the distribution of the risk score was presented in the heat map. CONCLUSION: We identified a novel seven-gene signature in patients with glioma, which could be used as a predictor for the prognosis of patients with glioma in the future.

Isodon eriocalyx and its bioactive component Eriocalyxin B enhance cytotoxic and apoptotic effects of gemcitabine in pancreatic cancer.

BACKGROUND: Pancreatic cancer, associated with poor prognosis and low survival rate, has been the fourth leading cause of cancer-related death in the US. Although gemcitabine (Gem) is the first-line chemotherapeutic drug in the management of pancreatic cancer, the median survival extension is only 1.5 months, indicating unsatisfactory clinical results. Therefore, exploring agents that can enhance the anti-cancer activity of Gem would be an attractive strategy. PURPOSE: Our previous studies have demonstrated that eriocalyxin b (EriB), an ent­kaurane diterpenoid isolated from Isodon eriocalyx (Dunn.) Hara, possesses anti-pancreatic cancer effects, thus acting as a potential therapeutic agent. In this study, we further investigated whether EriB or the ethanol extract of I. eriocalyx (Isodon) could potentiate the cytotoxic activity of Gem in human pancreatic adenocarcinoma cells. In addition, the mechanism associated with their effects was also studied. METHODS: The anti-proliferation effect was assessed by MTT assay and Ki-67 immunostaining. The combination effect (addition, synergism and antagonism) of various agents was calculated by the Calcusyn software (Biosoft), utilizing the T.C. Chou Method. Apoptosis was detected using Annexin V and PI double staining followed by quantitative flow cytometry. Protein expression regulated by various treatments was analyzed by western blotting. RESULTS: The combination index revealed that Gem and EriB (or Isodon extract) had synergistic anti-proliferative effect. Both cellular apoptotic and anti-proliferative effects of Gem were significantly increased after combination with EriB (or Isodon extract). The underlying mechanisms involved in the combination effects were elucidated, which include: (1) increased activation of the caspase cascade; (2) reduction of PDK1 and AKT phosphorylation; (3) induction of JNK phosphorylation by Isodon and Gem combination. CONCLUSION: Gem and EriB (or Isodon extract) taken together in combination regulated PDK1/AKT1/caspase and JNK signaling and promoted apoptosis synergistically, which may contribute to the much increased anti-proliferative activity compared to either agent alone.

The Effects of Lentivirus-mediated shRNA Interference Targeting Mcl-1 on Growth of NK/T-cell Lymphoma.

Myeloid leukemia-1 (Mcl-1) gene has been reported as an important factor in various types of cancer, but little research was processed on natural killer (NK)/T-cell lymphoma, a kind of a highly aggressive disease with a poor prognosis. Here we investigated the expression of Mcl-1 in seven lymphoma cell lines and its potential role as a molecular drug target for NK/T-cell lymphoma therapy by using lentivirus-mediated shRNA interference targeting Mcl-1 (lenti-shMcl-1). In our study, the expression of Mcl-1 in different lymphoma cell lines were evaluated firstly, after that lenti-shMcl-1 was constructed and transduced into NK/T-cell lymphoma cell line SNK-6 which had a high level expression of Mcl-1. Methylthiazolete-Trazolium (MTT) assay and flow cytometry (FCA) were employed to detect the status of proliferation and apoptosis after infection. Lastly we investigated the effects of chemotherapy agent vincristine (VCR) combination with lenti-shMcl-1 by MTT and FCA assay. The results showed that Mcl-1 gene expressed in all seven lymphoma cell lines at different levels. Recombinant lentiviruses could infect SNK-6 cells effectively. Mcl-1 expression level was remarkably down-regulated after infection with lenti-shMcl-1. The growth of SNK-6 cells was inhibited significantly through apoptosis pathway. Otherwise, lenti-shMcl-1 also revealed a significant chemosensitizing effect in combination with vincristine. In a word, we demonstrated that lenti-shMcl-1 had a significant anti-NK/T cell lymphoma effect and targeting Mcl-1 therapy could be a promising novel approach in treatment of lymphoma.

Eriocalyxin B-induced apoptosis in pancreatic adenocarcinoma cells through thiol-containing antioxidant systems and downstream signalling pathways.

Thiol-containing antioxidant systems play an important role in regulating cellular redox homeostasis. Several anti-cancer agents act by targeting these systems by inducing the production of reactive oxygen species (ROS). Our earlier studies have shown that Eriocalyxin B (EriB), a diterpenoid isolated from Isodon eriocalyx, possesses anti-pancreatic tumour activities in vitro and in vivo. The present study further demonstrated that only thiol-containing antioxidants, N-acetylcysteine (NAC) or dithiothreitol (DTT), inhibited EriB-induced cytotoxicity and apoptosis. EriB suppressed the glutathione and thioredoxin antioxidant systems, thus increasing the intracellular ROS levels and regulating the MAPK, NFκB pathways. Treatment with EriB depleted the intracellular thiol-containing proteins in CAPAN-2 cells. In vivo studies also showed that EriB treatment (2.5 mg/kg) reduced the pancreatic tumour weights significantly in nude mice with increased superoxide levels. Taken together, our results shed important new light on the molecular mechanisms of EriB acting as an apoptogenic agent and its therapeutic potential for pancreatic cancer.

Differential effect of MMSET mRNA levels on survival to first-line FOLFOX and second-line docetaxel in gastric cancer.

BACKGROUND: Breast cancer susceptibility gene 1 (BRCA1) expression differentially affects outcome to platinum- and taxane-based chemotherapy. Mediator of DNA damage checkpoint protein 1 (MDC1), p53-binding protein 1 (53BP1), multiple myeloma SET domain (MMSET) and ubiquitin-conjugating enzyme 9 (UBC9) are involved in DNA repair and could modify the BRCA1 predictive model. METHODS: Mediator of DNA damage checkpoint protein 1, 53BP1, MMSET and UBC9 mRNA were assessed in gastric tumours from patients in whom BRCA1 levels had previously been determined. RESULTS: In vitro chemosensitivity assay, MMSET levels were higher in docetaxel-sensitive samples. In a separate cohort, survival was longer in those with low MMSET (12.3 vs 8.8 months; P=0.04) or UBC9 (12.4 vs 8.8 months; P=0.01) in patients receiving only folinic acid, fluorouracil (5-FU) and oxaliplatin (FOLFOX). Conversely, among patients receiving second-line docetaxel, longer survival was associated with high MMSET (19.1 vs 13.9 months; P=0.003). Patients with high MMSET and BRCA1 attained a median survival of 36.6 months, compared with 13.9 months for those with high BRCA1 and low MMSET (P=0.003). In the multivariate analyses, low MMSET (hazard ratio (HR), 0.59; P=0.04) and low UBC9 (HR, 0.52; P=0.01) levels were markers of longer survival to first-line FOLFOX, whereas palliative surgery (HR, 2.47; P=0.005), low BRCA1 (HR, 3.17; P=0.001) and low MMSET (HR, 2.52; P=0.004) levels were markers of shorter survival to second-line docetaxel. CONCLUSIONS: Breast cancer susceptibility gene 1, MMSET and UBC9 can be useful for customising chemotherapy in gastric cancer patients.

Finite-size effect on magnetic properties in iron sulfide nanowire arrays.

We report the size effect on the magnetic properties in Fe(7)S(8) nanowire arrays. Samples with diameters in the range of 50-200 nm have been prepared by electrodeposition with AAO films. The Mössbauer measurement results show that four parameters (hyperfine fields, isomer shift, quadrupole splitting, full width at half-maximum) increased with decreasing the diameter of the nanowires. The magnetic properties were investigated. The hysteresis loop shape and the magnetization are dependent on the diameter of the nanowires. The thermomagnetic measurements on the as-synthesized nanowire samples and the corresponding bulk display a mixed-type curve and a Weiss-type curve, respectively.

Role of Nedd4-2 and polyubiquitination in epithelial sodium channel degradation in untransfected renal A6 cells expressing endogenous ENaC subunits.

Amiloride-sensitive epithelial sodium channels (ENaC) are responsible for transepithelial Na(+) transport in the kidney, lung, and colon. The channel consists of three subunits (alpha, beta, and gamma). In Madin-Darby canine kidney (MDCK) cells and Xenopus laevis oocytes transfected with all three ENaC subunits, neural precursor cell-expressed developmentally downregulated protein (Nedd4-2) promotes ubiquitin conjugation of ENaC. For native proteins in some cells, ubiquitin conjugation is a signal for their degradation by the ubiquitin-proteasome pathway, whereas in other cell types ubiquitin conjugation is a signal for endocytosis and lysosomal protein degradation. When ENaC are transfected into MDCK cells, ubiquitin conjugation leads to lysosomal degradation. In this paper, we characterize the involvement of the ubiquitin-proteasome proteolytic pathway in the regulation of functional ENaC in untransfected renal A6 cells expressing native ENaC subunits. In contrast to transfected cells, we show that total cellular alpha-, beta-, and gamma-ENaC subunits are polyubiquitinated and that ubiquitin conjugation of subunits increases when the cells are treated with a proteasome inhibitor. We show that Nedd4-2 is associated with alpha- and beta-subunits and is associated with the apical membrane. We also show the Nedd4-2 can regulate the number of functional ENaC subunits in the apical membrane. The results reported here suggest that the ubiquitin-proteasome proteolytic pathway is an important determinant of ENaC function in untransfected renal cells expressing endogenous ENaC.

Phosphatase inhibitors increase the open probability of ENaC in A6 cells.

We studied the cellular phosphatase inhibitors okadaic acid (OKA), calyculin A, and microcystin on the epithelial sodium channel (ENaC) in A6 renal cells. OKA increased the amiloride-sensitive current after approximately 30 min with maximal stimulation at 1-2 h. Fluctuation analysis of cell-attached patches containing a large number of ENaC yielded power spectra with corner frequencies in untreated cells almost two times as large as in cells pretreated for 30 min with OKA, implying an increase in single channel open probability (P(o)) that doubled after OKA. Single channel analysis showed that, in cells pretreated with OKA, P(o) and mean open time approximately doubled. Two other phosphatase inhibitors, calyculin A and microcystin, had similar effects on P(o) and mean open time. An analog of OKA, okadaone, that does not inhibit phosphatases had no effect. Pretreatment with 10 nM OKA, which blocks protein phosphatase 2A (PP2A) but not PP1 in mammalian cells, had no effect even though both phosphatases are present in A6 cells. Several proteins were differentially phosphorylated after OKA, but ENaC subunit phosphorylation did not increase. We conclude that, in A6 cells, there is an OKA-sensitive phosphatase that suppresses ENaC activity by altering the phosphorylation of a regulatory molecule associated with the channel.

Stimulation of chloride secretion by baicalein in isolated rat distal colon.

The effect of baicalein on mucosal ion transport in the rat distal colon was investigated in Ussing chambers. Mucosal addition of baicalein (1-100 microM) elicited a concentration-dependent short-circuit current (I(sc)) response. The increase in I(sc) was mainly due to Cl(-) secretion. The presence of mucosal indomethacin (10 microM) significantly reduced both the basal and subsequent baicalein-evoked I(sc) responses. The baicalein-induced I(sc) were inhibited by mucosal application of diphenylamine-2-carboxylic acid (100 microM) and glibenclamide (500 microM) and basolateral application of chromanol 293B (30 microM), a blocker of K(v)LQT1 channels and Ba(2+) ions (5 mM). Treatment of the colonic mucosa with baicalein elicited a threefold increase in cAMP production. Pretreating the colonic mucosa with carbachol (100 microM, serosal) but not thapsigargin (1 microM, both sides) abolished the baicalein-induced I(sc). Addition of baicalein subsequent to forskolin induced a further increase in I(sc). These results indicate that the baicalein evoked Cl(-) secretion across rat colonic mucosa, possibly via a cAMP-dependent pathway. However, the action of baicalein cannot be solely explained by its cAMP-elevating effect. Baicalein may stimulate Cl(-) secretion via a cAMP-independent pathway or have a direct effect on cystic fibrosis transmembrane conductance regulator.

Colon epithelial cell death in 2,4,6-trinitrobenzenesulfonic acid-induced colitis is associated with increased inducible nitric-oxide synthase expression and peroxynitrite production.

Peroxynitrite, derived from the reaction of nitric oxide (NO(.)) with superoxide (O(2)), is a potent nitrating and oxidizing agent that can induce apoptosis in a variety of different cell types. In the present study, we investigated the possible role of peroxynitrite as a mediator of colon epithelial cell death in rat colitis. Rat colon inflammation was induced by intracolonic administration of 2,4,6-trinitrobenzenesulfonic acid (TNBS) and rats were sacrificed 24 h after TNBS administration. Expression of inducible nitric-oxide synthase (iNOS) was detected by reverse transcription-polymerase chain reaction and immunohistochemistry. The enzymatic activities of Ca(2+)-independent iNOS and Ca(2+)-dependent constitutive nitric-oxide synthase were determined biochemically. Evidence of peroxynitrite-mediated cell injury was detected by immunostaining of nitrotyrosine. Apoptosis was examined by in situ terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay and DNA gel electrophoresis. To evaluate the specific contribution of peroxynitrite to the observed cell injury, a selective iNOS inhibitor, L-N(G)-[1-iminoethyl]lysine (L-NIL), was administered after TNBS induction. Morphological examination and analysis of TUNEL/cytokeratin double immunofluorescence revealed significant apoptosis in mucosal epithelial cells. Nitrotyrosine was colocalized with TUNEL, strongly demonstrating the association of peroxynitrite with the apoptotic death of colon epithelial cells. The administration of L-NIL reduced iNOS activity in 24-h lesions by 92% and also significantly attenuated both nitrotyrosine staining and apoptotic cell counts in the colon epithelium. These results strongly suggest that local elevated level of peroxynitrite produced from increased iNOS expression and activity is a major contributor to colon epithelial apoptosis during colon inflammation.

Enac degradation in A6 cells by the ubiquitin-proteosome proteolytic pathway.

Amiloride-sensitive epithelial Na(+) channels (ENaC) are responsible for trans-epithelial Na(+) transport in the kidney, lung, and colon. The channel consists of three subunits (alpha, beta, gamma) each containing a proline rich region (PPXY) in their carboxyl-terminal end. Mutations in this PPXY domain cause Liddle's syndrome, an autosomal dominant, salt-sensitive hypertension, by preventing the channel's interactions with the ubiquitin ligase Neural precursor cell-expressed developmentally down-regulated protein (Nedd4). It is postulated that this results in defective endocytosis and lysosomal degradation of ENaC leading to an increase in ENaC activity. To show the pathway that degrades ENaC in epithelial cells that express functioning ENaC channels, we used inhibitors of the proteosome and measured sodium channel activity. We found that the inhibitor, MG-132, increases amiloride-sensitive trans-epithelial current in Xenopus distal nephron A6 cells. There also is an increase of total cellular as well as membrane-associated ENaC subunit molecules by Western blotting. MG-132-treated cells also have increased channel density in patch clamp experiments. Inhibitors of lysosomal function did not reproduce these findings. Our results suggest that in native renal cells the proteosomal pathway is an important regulator of ENaC function.

Cryptdin-3 induces novel apical conductance(s) in Cl- secretory, including cystic fibrosis, epithelia.

Opening of anion-conductive pathways in apical membranes of secretory cells lining mucosal surfaces is a critical step in salt and water secretion and, thus, hydration of sites including airway and intestine. In intestine, Paneth cells are positioned at the base of the secretory gland (crypt) and release defensin peptide, in mice termed cryptdins, into the crypt lumen. Because at least some defensins have been shown to form anion-conductive channels in phospholipid bilayers, we tested whether these endogenous antimicrobial peptides could act as soluble inducers of channel-like activity when applied to apical membranes. To directly evaluate the possibility of cryptdin-3-mediated apical anion conductance (G(ap)), we have utilized amphotericin B to selectively permeabilize basolateral membranes of electrically tight monolayers of polarized human intestinal secretory epithelia (T84 cells), thus isolating the apical membrane for study. Cryptdin-3 induces G(ap) that is voltage independent (deltaG(ap) = 1.90 +/- 0.60 mS/cm2) and exhibits ion selectivity contrasting to that elicited by forskolin or thapsigargin (for cryptdin-3, Cl- = gluconate; for forskolin and thapsigargin, Cl- >> gluconate). We cannot exclude the possibility that the macroscopic current induced by cryptdin could be the sum of cation and Cl- currents. Cryptdin-3 induces a current in basolaterally permeabilized epithelial monolayers derived from airway cells harboring the deltaF508 mutation of cystic fibrosis (CF; deltaG(ap) = 0.80 +/- 0.06 mS/cm2), demonstrating that cryptdin-3 restores anion secretion in CF cells; this occurs independently of the CF transmembrane conductance regulator channel. These results support the idea that cryptdin-3 may associate with apical membranes of Cl--secreting epithelia and self-assemble into conducting channels capable of mediating a physiological response.

Mechanisms of aldosterone's action on epithelial Na + transport.

Aldosterone maintains total organism sodium balance in all higher vertebrates. The level of sodium reabsorption is primarily determined by the action of aldosterone on epithelial sodium channels (ENaC) in the distal nephron. Recent work shows that, in an aldosterone-sensitive renal cell line (A6), aldosterone regulates sodium reabsorption by short- and long-term processes. In the short term, aldosterone regulates sodium transport by inducing expression of the small G-protein, K-Ras2A, by stimulating the activity of methyl transferase and S-adenosyl-homocysteine hydrolase to activate Ras by methylation, and, possibly, by subsequent activation by K-Ras2A of phosphatidylinositol phosphate-5-kinase (PIP-5-K) and phosphatidylinositol-3-kinase (PI-3-K), which ultimately activates ENaC. In the long term, aldosterone regulates sodium transport by altering trafficking, assembly, and degradation of ENaC.

Aldosterone induces ras methylation in A6 epithelia.

Aldosterone increases Na(+) reabsorption by renal epithelial cells: the acute actions (<4 h) appear to be promoted by protein methylation. This paper describes the relationship between protein methylation and aldosterone's action and describes aldosterone-mediated targets for methylation in cultured renal cells (A6). Aldosterone increases protein methylation from 7.90 +/- 0.60 to 20.1 +/- 0.80 methyl ester cpm/microg protein. Aldosterone stimulates protein methylation by increasing methyltransferase activity from 14.0 +/- 0.64 in aldosterone-depleted cells to 31.8 +/- 2.60 methyl ester cpm/microg protein per hour in aldosterone-treated cells. Three known methyltransferase inhibitors reduce the aldosterone-induced increase in methyltransferase activity. One of these inhibitors, the isoprenyl-cysteine methyltransferase-specific inhibitor, S-trans, trans-farnesylthiosalicylic acid, completely blocks aldosterone-induced protein methylation and also aldosterone-induced short-circuit current. Aldosterone induces protein methylation in two molecular weight ranges: near 90 kDa and around 20 kDa. The lower molecular weight range is the weight of small G proteins, and aldosterone does increase both Ras protein 1.6-fold and Ras methylation almost 12-fold. Also, Ras antisense oligonucleotides reduce the activity of Na(+) channels by about fivefold. We conclude that 1) protein methylation is essential for aldosterone-induced increases in Na(+) transport; 2) one target for methylation is p21(ras); and 3) inhibition of Ras expression or Ras methylation inhibits Na(+) channel activity.

Measurement of tissue electrical impedance confirms stereotactically localized internal segment of the globus pallidus during surgery.

Lesions surgically made in the internal segment of the posteroventral globus pallidus (Gpi) reduce many medically intractable symptoms in patients with Parkinson's disease. The Gpi is localized for pallidotomy by stereotactic procedures. We sought to confirm the stereotactically localized Gpi segment by measuring impedance between the tip of an active electrode that was advanced toward the target and a remote reference electrode. A stereotactic instrument was used to place an active electrode in 53 conscious patients undergoing pallidotomy. The electrode was manually advanced slowly toward the Gpi segment by a microdrive. Impedance was measured every 10 mm initially and every 1 mm in the final 10 mm of the electrode's advancement, using an impedance monitor interconnected with a lesion generator. Measurements were continued for several millimeters after each patient's target was reached. Impedance values ranged from 250 to 350 ohms throughout of the electrode passage. Impedance increased by 25-35 ohms at 1-3 mm before the center of the target and then dropped abruptly when the electrode passed the target. Impedance values depended strongly on the electrode's configuration. The impedance measurement confirmed the location of each surgical target identified by stereotactic navigation. The accuracy of the impedance measurement is acceptable, and the procedure can be recommended as an aid to confirm the stereotactically localized Gpi during neurological surgery.

Regulation of Na(+) reabsorption by the aldosterone-induced small G protein K-Ras2A.

Xenopus laevis A6 cells were used as model epithelia to test the hypothesis that K-Ras2A is an aldosterone-induced protein necessary for steroid-regulated Na(+) transport. The possibility that increased K-Ras2A alone is sufficient to mimic aldosterone action on Na(+) transport also was tested. Aldosterone treatment increased K-Ras2A protein expression 2.8-fold within 4 h. Active Ras is membrane associated. After aldosterone treatment, 75% of K-Ras was localized to the plasma membrane compared with 25% in the absence of steroid. Aldosterone also increased the amount of active (phosphorylated) mitogen-activated protein kinase kinase likely through K-Ras2A signaling. Steroid-induced K-Ras2A protein levels and Na(+) transport were decreased with antisense K-ras2A oligonucleotides, showing that K-Ras2A is necessary for the natriferic actions of aldosterone. Aldosterone-induced Na(+) channel activity, was decreased from 0.40 to 0.09 by pretreatment with antisense ras oligonucleotide, implicating the luminal Na(+) channel as one final effector of Ras signaling. Overexpression of K-Ras2A increased Na(+) transport approximately 2.2-fold in the absence of aldosterone. These results suggest that aldosterone signals to the luminal Na(+) channel via multiple pathways and that K-Ras2A levels are limiting for a portion of the aldosterone-sensitive Na(+) transport.

Role of interferon-gamma in lung inflammation following cecal ligation and puncture in rats.

Interferon-gamma (IFN-gamma) has been implicated in the mortality of animal models of endotoxemia. On the other hand, the specific role of IFN-gamma in the development of organ inflammation in a model of polymicrobial sepsis has not been elucidated. In this study, we hypothesized that IFN-gamma plays an important role in lung inflammation after cecal ligation and puncture (CLP). To verify this hypothesis, lung tissue was removed 5 h after CLP or from sham controls. The mRNA expression (by RT-PCR) of IFN-gamma was increased in lung homogenates of CLP rats compared to sham controls. Using immunohistochemistry, we show for the first time the increased presence of IFN-gamma staining cells in the lung following CLP. Only very small amounts of positive staining for IFN-gamma was observed in lungs of sham controls. The presence of IFN-gamma in the lung 5 h after CLP correlated with a twofold increases in lung superoxide generation and MPO activity (index of neutrophil sequestration). Plasma and lung nitrite levels (breakdown product of nitric oxide) were also significantly increased in CLP rats. IFN-gamma antibody (1.2 mg/kg, i.v.) administered immediately after CLP significantly decreased lung superoxide levels to levels similar to the sham controls without affecting MPO activity, or lung or plasma nitrite levels. These results provide evidence that IFN-gamma may contribute to lung inflammation 5 h following CLP via increased production of superoxide.

Measurements of the neutron yields from 7Li(p,n)7Be reaction (thick target) with incident energies from 1.885 to 2.0 MeV.

Accelerator-based neutron source have been considered to be practical for boron neutron capture therapy (BNCT). Based on experience with a parameters of the Brookhaven National Laboratory BMRR reactor neutron source, which has been used in treatment experiments, the future accelerator-based neutron source for BNCT should have the properties of low energy distribution (< 100 keV) and high flux (about 10(9) neutrons per second per square centimeter) in the patient zone. Using protons to bombard thick 7Li targets, generating neutrons via the 7Li(p,n)7Be reaction, is one of the optimal choices for this kind of neutron source. Neutron yield data versus incident energy are necessary in order to select the proper incident energy and for estimating how high the incident proton current should be. The required proton beam current intensity is one of the key parameters for an accelerator useful for BNCT. In the present work, neutron yields of the 7Li(p,n)7Be reaction with a thick lithium target and incident energies of 1.885 and 1.9 MeV were measured at 0 degree with respect to the incident beam direction. The results are (3.08 +/- 0.17) x 10(12) and (5.71 +/- 0.32) x 10(12) neutrons/C sr, respectively. Neutron yield angular distribution measurements at 2 MeV incident energy were also performed. The proton beams were generated by the Peking University 4.5 MV electrostatic accelerator. The emitted neutrons from these reactions have the advantages of low energy distribution and forward angular distribution, which are requirements for a BNCT neutron source. The data obtained in this work can be used as a reference to study the accelerator-based neutron sources for BNCT.