Dynamics of isoproterenol subsensitivity in guinea pig airway smooth muscle

Possible subsensitivity to the bronchodilator effects of beta adrenergic agonists has been a clinical concern for the past decade. We have examined this possibility in guinea pigs who were treated in vivo with isoproterenol, either 375 µg/kg BW (single dose) or 75 µg/kg BW every 20 min for 5 h (multiple dose). Tracheal segments were isolated from control and treated animals at 1/2, 1, 2, 4, and 6 h after the last injection and the isoproterenol cumulative dose-response relaxation curves were examined in vitro (n=92 prep.). Segments of the same tracheas were also used for measurement of “basal” cAMP and cAMP response to isoproterenol stimulation in vitro (n=78 prep.). Subsensitivity to isoproterenol-induced tracheal relaxation was present 1 h after single or multiple doses of in vivo treatment. At all other times the ED50 for tracheal relaxation was similar to the control group. There were marked alterations in tracheal cAMP responses to adrenergic stimulation in vitro. After single doses of in vivo treatment, cAMP response was markedly enhanced at 1/2 h, suppressed at 1 h, and normal thereafter. After multiple doses, cAMP was suppressed for 6 h. This difference in the duration of subsensitivity in the two systems suggests that the beta adrenergic effects on airway smooth muscle may be mediated through mechanisms other than tissue cAMP levels.     

Accumulation of histamine and cyclic nucleotides in lung tissue during chronic hypobaric hypoxia

Rats exposed to hypobaric hypoxia (P_B = 440 Torr) from 1 to 9 days showed a rise of the total lung histamine content which reached a maximum of 1.138 ng/µg DNA = 274%; (control values were 0.415 ng/µg DNA = 100%) after 2 days of hypoxia. The cyclic AMP (cAMP) concentrations were elevated during the first 3 days of hypobaric hypoxia and returned almost to control values later on in the time course. Cyclic GMP (cGMP) increased after an initial delay, reaching plateau values from the 6th to the 9th day. Histamine and cAMP concentrations were highly correlated during the first 3 days of hypoxia. 6-Methylprednisolone increased lung tissue concentrations of both histamine and cAMP in hypoxic as well as nonhypoxic animals; the level of cGMP was unchanged. The results demonstrate an accumulation of histamine in lung tissue and suggest that histamine is stored rather than released from rat lungs during chronic hypoxia. The possible role of cyclic nucleotides in the regulation of histamine lung tissue content is discussed.     

The significance of cyclic 3′5′-AMP for the euler-liljestrand mechanism

Changes in the pulmonary vascular resistance and the cAMP levels in the lung parenchyma of four pigs were recorded during normoxia and hypoxia and following the administration of orciprenaline under hypoxia. Additional measurements recorded were pulmonary artery pressure, left-ventricular pressure, dp/dt_max, mean arterial pressure, arterial oxygen saturation, blood gas values. The rise of the pulmonary vascular resistance during hypoxia is not preceded by a decrease in intracellular cAMP levels; extreme cAMP concentrations, however, block the Euler-Liljestrand mechanism. The findings suggest that alveolar hypoxia and/or intracellular metabolic acidosis suppresses the inhibiting action of cAMP on the Ca⁺⁺ influx in the smooth vascular muscles or affect the Ca⁺⁺-binding process of the sarcoplasmic reticulum of the vascular musculature, resulting in vascoconstriction.     

Fecal fistula: A late complication of Marlex® mesh repair

Marlex^® mesh is an excellent prosthetic material for closure of major abdominal defects. Most of its complications are seroma and infections. We have used Marlex mesh intraperitoneally for closure of burst abdomen in a patient who had a gastrectomy for recurrent duodenal ulcer. A year and a half later, this patient developed a fecal fistula to the skin due to incorporation of the Marlex mesh into the splenic flexure of the colon. The patient underwent a second operation during which the fistula was resected and the Marlex removed. We concluded that intraperitoneal placement of Marlex mesh is not recommended.     

Prostaglandin E in cholera toxin-induced intestinal secretion

Prostaglandin E₁ (PGE₁) and cholera enterotoxin stimulate small-intestine mucosal adenylate cyclase and intestinal secretion of water and electrolytes. The previous suggestion that PGE may mediate cholera-toxin effects was explored in these studies. Closed rabbit jejunal loops were injectedin vivo with cholera toxin and compared to similar loops in the same animal injected with buffer. Loop mucosal homogenates and intestinal secretions were analyzed by radioimmunoassay for cAMP and PGE concentrations. Cholera toxin produced significant increases in mucosal and intestinal fluid cAMP; however, there were no significant increases in PGE in the toxin-treated loops when compared to the control loops. In addition, there was no correlation between cAMP and PGE in the same samples. These studies indicate that cholera toxin stimulates intestinal cAMP and secretion independent of PGE synthesis and provide evidence against a specific role for PGE in mediating cholera-toxin effects.     

Comparative study of free omental,peritonenal, Dacron velour,and Marlex mesh reinforcement of large-bowel anstomosis

The comparative role of free ometal, peritoneal, Dacron^® velour, and Marlex^® mesh grafts in reinforcement of an extremely vulnerable experimental model of large-bowel anastomosis was studied in dogs. While both the omentum and peritoneum proved not to be effective in preventing anastomotic leakage, Dacron velour did considerably lower this incidence to within reasonable limits but led to formation of low-grade lymphoma at the reinforcement site in two animals. Only Marlex mesh was found to be highly effective in sealing the suture line, and it is anticipated that, with the usual teachnique of anastomosis, this sealing effect will be foolproof, thereby nullifying any risk of suture-line breakdown     

Piperlongumine induces cell death through ROS-mediated CHOP activation and potentiates TRAIL-induced cell death in breast cancer cells

Purpose

Piperlongumine (PL) has been shown to selectively induce apoptotic cell death in cancer cells via reactive oxygen species (ROS) accumulation. In this study, we characterized a molecular mechanism for PL-induced cell death.

Methods

Cell viability and cell death were assessed by MTT assay and Annexin V-FITC/PI staining, respectively. ROS generation was measured using the H₂DCFDA. Small interfering RNA (siRNA) was used for suppressing gene expression. The mRNA and protein expression were analyzed by RT-PCR and Western blot analysis, respectively.

Results

We found that PL promotes C/EBP homologous protein (CHOP) induction, which leads to the up-regulation of its targets Bim and DR5. Pretreatment with the ROS scavenger N-acetyl-cysteine abolishes the PL-induced up-regulation of CHOP and its target genes, suggesting an essential role for ROS in PL-induced CHOP activation. The down-regulation of CHOP or Bim with siRNA efficiently attenuates PL-induced cell death, suggesting a critical role for CHOP in this cell death. Furthermore, PL potentiates TRAIL-induced cytotoxicity in breast cancer cells by upregulating DR5, as DR5 knockdown abolished the sensitizing effect of PL on TRAIL responses.

Conclusions

Overall, our data suggest a new mechanism for the PL-induced cell death in which ROS mediates CHOP activation, and combination treatment with PL and TRAIL could be a potential strategy for breast cancer therapy.     

Cell mediated immunity and suppressor T cell function in children with cystic fibrosis

The relationship of general and suppressor T cell function to IgE sensitization was investigated in a group of 12 children with cystic fibrosis (CF) and 10 non-atopic control children. Increased IgE sensitization was noted in the cystic fibrosis group, as measured by increased group mean serum IgE levels and an increased incidence of multiple IgE skin sensitivities. Studies of active E rosettes, lymphocyte stimulation, delayed-type skin responsiveness, and Concanavalin A (Con A) induced suppressor cell function revealed no statistically significant group differences. The results of this study may implicate multiple mechanisms for IgE sensitization.     

Serum and intestinal fluid immunoglobulins in patients with giardiasis

To test the possibility that a factor predisposing to symptomatic giardiasis in the general population is immunoglobulin deficiency, we measured immunoglobulins in serum and intestinal fluids of 9 giardiasis patients (8 after eradication ofGiardia lamblia) and 12 healthy individuals. The concentrations of IgG, IgA, IgM, and IgE in serum and of IgA and IgM in intestinal fluids from the 2 groups were similar. Intestinal fluid IgG levels were significantly higher (P<0.1), however, in the patients (5.8±6.3 mg/100 ml) than in the control subjects (1.1±1.9 mg/100 ml). This difference was unexplained and is of uncertain biological significance. We conclude that giardiasis in generally healthy individuals is not etiologically related to unsuspected immunoglobulin deficiencies, even thoughG. lamblia infection is known to be very common in grossly immunoglobulin-deficient patients.     

Mutations of EGFR or KRAS and expression of chemotherapy-related genes based on small biopsy samples in stage IIIB and IV inoperable non-small cell lung cancer

Purposes

Epidermal growth factor receptor (EGFR) and KRAS mutations may predict the outcome of targeted drug therapy and also may be associated with the efficacy of chemotherapy in patients with non-small cell lung cancer (NSCLC). This report investigated the relation of EGFR or KRAS mutation and expression of chemotherapy-related genes, including excision repair cross-complementing 1 (ERCC1), thymidylate synthetase (TYMS), ribonucleotide reductase subunit M1 (RRM1) and class III β-tubulin (TUBB3), as a potential explanation for these observations.

Methods

A total of 143 patients with stage IIIB and IV NSCLC from bronchoscopy or percutaneous lung biopsy obtained tumor samples were analyzed concurrently for EGFR or KRAS mutations, and mRNA expression of ERCC1, TYMS, RRM1 and TUBB3. EGFR or KRAS mutations were detected with xTAG liquidchip technology (xTAG-LCT), and mRNA expression levels of four genes were detected by branched DNA-liquidchip technology (bDNA-LCT).

Results

Of 143 patients, 63 tumors were positive for EGFR-activating mutations, and 16 tumors were positive for KRAS mutations. EGFR-activating mutations are more frequent in females, adenocarcinoma and non-smokers patients, and KRAS mutations are more frequent in smoking patients. ERCC1 mRNA levels were significantly associated with histological type and tumor differentiation, whereas TYMS levels were significantly associated with age. NSCLC specimens that harboring EGFR-activating mutations are more likely to express low ERCC1 and high TUBB3 mRNA levels, whereas tumors from patients with NSCLC harboring KRAS mutation are more likely to express high ERCC1 mRNA levels.

Conclusions

Mutations and expression of chemotherapy-related genes may provide a basis for the selection of suitable molecular markers for individual treatment in a population with stage IIIB and IV NSCLC.     

The BRAF mutation is associated with the prognosis in colorectal cancer

Background

Two members of the Ras/Raf signaling pathway, KRAS and B-raf, are suspected to be involved in the stepwise progression of colorectal cancer (CRC) tumorigenesis.

Objective

We compared the KRAS and BRAF mutation status of CRC patients with their clinicopathological characteristics and examined the effect of mutation status on survival rates.

Methods

DNA was extracted from 164 samples, and the mutation statuses of KRAS and BRAF were assessed using peptide PNA clamp real-time PCR method. The presences of mutation were compared with clinicopathological factors and 5-year survival rate.

Results

Among the 164 CRC cases, KRAS mutation as detected in 71 cases (43.3 %), respectively, with no relationship with clinicopathological factors of the patients. On Kaplan–Meier survival analysis, KRAS mutation was not significantly associated with survival (p = 0.971). BRAF mutation was detected in 26 cases (15.9 %) and not associated with clinicopathological factors of the patients. However, the 5-year survival rate of BRAF mutations was significantly decreased (p = 0.02).

Conclusions

The presence of KRAS mutation did not correlate with the various clinicopathological factors of CRC patients or the survival rate. However, the survival rate was reduced in BRAF-mutated CRC patients. Therefore, BRAF mutation could be an important prognostic factor in CRC patients.     

Inhibition of pentagastrin-stimulated gastric secretion by duodenal acidification or administration of fat in normal subjects and in patients with duodenal ulcer

The inhibitory effect of duodenal acidification and intraduodenal fat infusion on pentagastrin-stimulated gastric secretion in normal subjects and in patients with duodenal ulcer was studied. Intraduodenal infusion of acid resulted in inhibition of HCl secretion found to be significant only in ulcer patients. Pepsin output, although lower during the first 15 minutes of duodenal acidification, later increased. Intraduodenal infusion of olive oil resulted in significant inhibition of HCl and pepsin output in both groups of patients, which was maximal 45–60 minutes after the beginning of fat infusion. Gastric secretion was more readily inhibited in ulcer patients than in normal subjects; this difference was particularly evident in inhibition of pepsin secretion. In addition, decrease in concentration of HCl and pepsin was observed to be significant only in ulcer patients. Mechanisms by which duodenal acidification and fat inhibit gastric secretion are discussed. The results obtained suggest that secretin, which is probably responsible for inhibition after duodenal acidification, is not the inhibitor during inhibition by fat. The ulcer patients were found to have unimpaired mechanisms of inhibition by acid and fat.     

The humoral immune system in inflammatory bowel disease

As there have been reports of differences in mean levels of serum immunoglobulins between patients with ulcerative colitis and Crohn's disease, serum IgG, IgA, and IgM were estimated in 158 patients with inflammatory bowel disease and the results correlated with the clinical features of the patients. Although a higher mean IgG level in ulcerative colitis compared to Crohn's disease was confirmed, no difference was found when the comparison was limited to patients with colonic Crohn's disease. Patients with either disease had higher mean IgM levels than controls, and the IgM levels were higher on treatment with corticosteroids and showed a tendency to rise in remission. IgG and IgM levels were also higher in both diseases if extraintestinal manifestations were present. It is concluded that if clinical features, particularly disease site, are taken into account, the overall immunoglobulin responses in these two diseases show no differences.     

Suppression of Placental Metallothionein 1 and Zinc Transporter 1 mRNA Expressions Contributes to Fetal Heart Malformations Caused by Maternal Zinc Deficiency

Zinc has been implicated to have a protective role against heart malformations during fetal development. Metallothionein 1 (MT-1) and zinc transporter 1 (ZnT-1) are two major metabolic factors that are associated with zinc metabolism. The present work aimed to investigate the association of placental MT-1 and ZnT-1 expressions with fetal heart malformations resulting from maternal zinc deficiency. Sprague–Dawley female rats were randomly divided into five groups of extremely low-zinc, low-zinc, moderately low-zinc, marginally low-zinc and normal zinc (n = 9–12), and were fed diets with controlled zinc content at 1.0 ± 0.3, 8.4 ± 1.8, 15.4 ± 2.8, 22.4 ± 4.1 and 29.4 ± 5.3 [mean ± standard deviation (SD)] mg of zinc/kg, respectively, from day 25 of preconception until day 19 of gestation. The female rats were bred, their fetuses were harvested at day 19 of gestation after killing the dams, and fetal hearts were morphologically examined. Zinc concentration and alkaline phosphatase (ALP) activity in maternal venous blood sera were tested, and MT-1 and ZnT-1 mRNA expressions in the placenta were assayed. Zinc concentrations and ALP activities in the blood were low in all zinc-deficient diet groups in a dose-dependent fashion. The incidences of heart malformations were increased, and the levels of placental MT-1 and ZnT-1 mRNA expressions were decreased in the extremely low-zinc, low-zinc and moderately low-zinc groups compared with the normal zinc group. Specifically, mRNA levels of placental MT-1 or ZnT-1 were significantly decreased and were lower than the specific threshold values in the fetuses with heart malformations but not in the fetuses without heart malformations in all the groups. These data indicate that maternal zinc deficiency resulted in an elevated incidence of fetal heart malformations, which was associated with significant decreases in placental MT-1 and ZnT-1 mRNA expressions to the levels below the threshold values that may be a crucial factor to determine the presence of fetal heart malformations.     

Associations between functional TNFR2 196 M/R polymorphisms and susceptibility to rheumatoid arthritis: a meta-analysis

Several studies have examined the effects of tumor necrosis factor receptor (TNFR) 1 +38 A/G and TNFR2 196 M/R polymorphisms on susceptibility to RA and have reported conflicting results. The purpose of this study was to examine whether the TNFR1 +38 A/G and TNFR2 196 M/R polymorphisms are associated with RA susceptibility. We performed a literature search using the Medical Literature Analysis and Retrieval System Online and Embase citation indices, and conducted a meta-analysis to examine the association between the TNFR1 +38 A/G and TNFR2 196 M/R polymorphisms and RA. Our meta-analysis included a total of 13 studies from 11 articles, consisting of 11 studies of the TNFR2 polymorphism (2,092 cases and 1,483 controls), and two studies of the TNFR1 polymorphism (672 cases and 288 controls). The meta-analysis revealed a significant association between the TNFR2 196 RR genotype and RA risk (OR 1.737, 95 % CI 1.275–2.367, P = 4.6 × 10^?5). Stratification by ethnicity indicated an association between the TNFR2 196 RR genotype and RA in Europeans (OR 2.054, 95 % CI 1.305–3.232, P = 0.002), but not in East Asians (OR 1.596, 95 % CI 0.642–3.971, P = 0.314). Analysis using a homozygote contrast model showed the same pattern for the TNFR2 196 RR genotype in a European and East Asian population. However, no association was found between the TNFR1 +36 A/G polymorphism and RA in a European population. Our meta-analysis demonstrated that the functional TNFR2 196 M/R polymorphism is associated with susceptibility to RA in the European population.     

Acyl CoA synthetase from rat lung purification and properties

Long chain fatty acyl-CoA synthetase, a key enzyme in fatty acid metabolism, occurs in liver, brain, muscle, adipose tissue and bacteria. In our study, rat lung was found to have an acyl-CoA synthetase very active for the synthesis of palmitoyl CoA. The lung microsomal and cell membrane fractions were the principle sources of this enzyme, with mitochondria the next most active fraction. Palmitoyl CoA synthetase was isolated from rat lung microsomes and purified 100-fold. Enzyme activity was determined either by spectrophotometric or by radioactive assay methods. Kinetic parameters and properties were determined using purified microsomal palmitoyl CoA synthetase. The assay system required ATP and CoA as substrates. Maximal activation was reached with palmitate as substrate.     

AIDS prevention in primary care clinics

To assess attitudes toward educational programs about AIDS, 540 patients and 36 of their medical providers in primary care clinics were systematically sampled to ascertain what age groups should be exposed to a pamphlet entitled “Am I at Risk for AIDS?”, as well as what was acceptable content for posters and pamphlets placed in clinic waiting rooms. Although fewer than 10% of patients and providers opposed asking both teenagers and adults to read a pamphlet listing risk groups and practices, 24% of patients and 51% of providers opposed exposing children to the pamphlets. Only 6% of patients and none of the providers opposed all posters about AIDS, but 30% of patients and 44% of providers opposed posters listing risk groups, and opposition was even greater to posters describing “safe sex.” Regarding pamphlets acceptable for clinic waiting rooms, resistance among patients and providers was common (>25%) only when sexual practices were specifically mentioned. Although 89% of patients stated they had never been asked about their sexual orientations by a doctor or nurse, 34% of providers stated that they “always” or “often” so inquire. In the primary care clinics surveyed, a wide variety of educational interventions about AIDS could be implemented with little opposition.     

Circulating immune complexes in Crohn's disease

Of the sera from 55 patients with Crohn's disease, 37% showed evidence of anticomplementary activity. C3 and C4 levels were significantly increased in these patients compared with control subjects, although total hemolytic complement levels were normal. C3 inactivation products were demonstrated in 32.9% of serum and plasma samples from 51 patients with CD. This was taken to be evidence ofin vivo activation of C3 and the complement system. Activation of the C′ system was thought to be caused by circulating immune complexes since evidence of anticomplementary activity could be correlated with incidence of C3 activation. Gel filtration studies carried out on 15 serum samples (13 patients) showing anticomplementary activity showed this activity to be confined to two fractions of molecular weight range >10⁶ and 2×10⁵?10⁶ daltons. IgG and IgA were present in both these fractions. As the molecular weights of IgG and IgA are lower than the molecular weight range of these fractions, it is possible that the immune complexes were composed of IgG and possibly IgA complexed with antigen.     

Biliary ascariasis diagnosed by ERCP with chronic dilation n of the biliary system

In February of 1970 when I was the gastroenterologist at the Oak Knoll Naval Hospital, I received a notice that Dr. Ingelfinger was coming to San Francisco to be a visiting professor. I decided that I would definitely go over to hear him and to see how things were back in Boston. The morning of the presentation, as I eatieating breakfast, I suddenly turned to my wife, Gail, and said, “What if he asks me a question?” Gail looked up at me and said, “Oh, Ralph, you're not a fellow anymore. Dr. Ingelfinger wouldn't do anything like that.” That was scant reassurance for me; nevertheless, I was at the meeting at 9:00 and had a brief moment to say hello to Dr. Ingelfinger before the case was presented to him. It was, in fact, a very difficult case of a 9-year-old girl with pancreatitis.As Dr. Ingelfinger went through the differential diagnosis of pancreatitis including hyperlipemia, hypercalcemia, and familial, he suddenly looked at me and said, “Ralph, what other diffential should we be thinking of?” I sat there dumbfounded. I couldn't believe that he was going to get me again. As I sat there he looked up in disgust and said, “Ascaris,Ralph, Ascaris. You've got to think ofAscaris worms in a young girl with pancreatitis.” I didn't get it that day, but here is a case of ascariasis for Dr. Ingelfinger.