The rates of second lung cancers and the survival of surgically-resected second primary lung cancers in patients undergoing resection of an initial primary lung cancer.

BACKGROUND: The Lung Cancer Screening Trial demonstrated improved overall survival (OS) and lung cancer specific survival (LCSS), likely due to finding early-stage NSCLC. The purpose of our investigation is to evaluate whether long-term surveillance strategies (4+ years after surgical resection of the initial lung cancer(1LC)) would be beneficial in NSCLC patients by assessing the rates of second lung cancers(2LC) and the OS/LCSS in patients undergoing definitive surgery in 1LC as compared to 2LC (>48 months after 1LC) populations. METHODS: SEER13/18 database was reviewed for patients during 1998-2013. Log-rank tests were used to determine the OS/LCSS differences between the 1LC and 2LC in the entire surgical group(EG) and in those having an early-stage resectable tumors (ESR, tumors <4 cm, node negative). Joinpoint analysis was used to determine rates of second cancers 4-10 year after 1LC using SEER-9 during years 1985-2014. RESULTS: The rate of 2LCs was significantly less than all other second cancers until 2001 when the incidence of 2LCs increased sharply and became significantly greater than all other second cancers in females starting in year 2005 and in men starting in year 2010. OS/LCSS, adjusted for propensity score by using inverse probability weighting, demonstrated similar OS, but worse LCSS for 2LCs in the EG, but similar OS/LCSSs in the ESR group. CONCLUSION: Because the rate of 2LCs are increasing and because the OS/LCSS of the 1LC and 2LC are similar in early-stage lesions, we feel that continued surveillance of patients in order to find early-stage disease may be beneficial.

Temporal migration of gonadotrophin-releasing hormone-1 neurones is modified in GAD67 knockout mice.

Gonadotrophin-releasing hormone (GnRH-1) neurones reside in the forebrain and regulate gonadal function via the hypothalamic-pituitary-gonadal axis. Disruption of this axis results in reproductive dysfunction. During embryonic development, GnRH-1 neurones migrate from the nasal pit through the nasal/forebrain junction (NFJ) into the developing brain. Prenatally gamma-aminobutyric acid (GABA) is excitatory and has been shown to play a role in nervous system development. Both in vivo and in vitro experiments suggest that GABA inhibits migration of GnRH-1 neurones. The present study examines the migration of GnRH-1 neurones in GAD67 knockout (KO) mice to further elucidate the role of GABA on GnRH-1 neuronal development. Three stages were examined, embryonic day (E)12.5, E14.5 and E17.5. GnRH-1 cell number and location were analysed by immunocytochemistry and in situ hybridisation histochemistry. The total number of GnRH-1 immunopositive cells was similar between wild-type (WT) and KO mice. However, significant differences were found in the overall distribution of GnRH-1 immunopositive cells in GAD67 KO compared to WT mice at all stages. Subsequent analysis by area revealed differences occurred at the NFJ with an increase in GnRH-1 cells in GAD67 KO at E14.5 and a decrease in GnRH-1 cells in GAD67 KO at E17.5. Comparable counts for cells expressing GnRH-1 transcript and protein were obtained. These data indicate that attenuated levels of GABA accelerate GnRH-1 cell migration in nasal areas as well as movement of GnRH-1 cells into the central nervous system at the NFJ.

Renal disease in rats with type 2 diabetes is associated with decreased renal nitric oxide production.

AIMS/HYPOTHESIS: In several other models of chronic renal disease, decreases in renal nitric oxide activity and nitric oxide synthase (NOS) protein abundance have been demonstrated. Here, we studied diabetic obese Zucker (ZDF Gmi fa/fa) rats that develop severe hyperglycaemia and renal disease, together with their lean control animals, to determine if renal nitric oxide deficiency also occurs in this model. METHODS: Obese Zucker rats aged 10 to 12 weeks were maintained on Purina 5008 diet until 4, 8, or 11 months of age and compared with similarly maintained, 4- and 11-month-old lean Zucker rats. NOS activity and abundance of endothelial NOS (eNOS) and neuronal NOS (nNOS) were measured on homogenates of kidney cortex. Blood was analysed for glucose, lipids, creatinine, and blood urea nitrogen and kidney tissue was obtained for histology. RESULTS: Obese rats exhibited severe hyperglycaemia from 4 months of age and developed increasing hyperlipidaemia, proteinuria, and decreasing renal function with age compared to lean counterparts. At 4 months cortical NOS activity and nNOS abundance were lower in obese rats than in lean ones. At 11 months NOS activity remained depressed and nNOS abundance had declined further in obese rats. Glomerulosclerosis in the obese rats was mild at 4 months, becoming severe by 11 months. Lean rats had only mild age-dependent increases in glomerular injury. CONCLUSIONS/INTERPRETATION: The chronic renal disease that occurs in hyperglycaemic, obese Zucker rats is associated with decreased renal cortical nitric oxide production and increasing renal injury, although the changes do not resemble those of diabetic nephropathy in man.

Dynamic expression of a glutamate decarboxylase gene in multiple non-neural tissues during mouse development.

BACKGROUND: Glutamate decarboxylase (GAD) is the biosynthetic enzyme for the neurotransmitter gamma-aminobutyric acid (GABA). Mouse embryos lacking the 67-kDa isoform of GAD (encoded by the Gad1 gene) develop a complete cleft of the secondary palate. This phenotype suggests that this gene may be involved in the normal development of tissues outside of the CNS. Although Gad1 expression in adult non-CNS tissues has been noted previously, no systematic analysis of its embryonic expression outside of the nervous system has been performed. The objective of this study was to define additional structures outside of the central nervous system that express Gad1, indicating those structures that may require its function for normal development. RESULTS: Our analysis detected the localized expression of Gad1 transcripts in several developing tissues in the mouse embryo from E9.0-E14.5. Tissues expressing Gad1 included the tail bud mesenchyme, the pharyngeal pouches and arches, the ectodermal placodes of the developing vibrissae, and the apical ectodermal ridge (AER), mesenchyme and ectoderm of the limb buds. CONCLUSIONS: Some of the sites of Gad1 expression are tissues that emit signals required for patterning and differentiation (AER, vibrissal placodes). Other sites correspond to proliferating stem cell populations that give rise to multiple differentiated tissues (tail bud mesenchyme, pharyngeal endoderm and mesenchyme). The dynamic expression of Gad1 in such tissues suggests a wider role for GABA signaling in development than was previously appreciated.

Intrarenal infusion of bradykinin elicits a pressor response in conscious rats via a B2-receptor mechanism.

Bradykinin (BK) is a peptide known to activate afferent nerve fibers from the kidney and elicit reflex changes in the cardiovascular system. The present study was specifically designed to test the hypothesis that bradykinin B2 receptors mediated the pressor responses elicited during intrarenal bradykinin administration. Pulsed Doppler flow probes were positioned around the left renal artery to measure renal blood flow (RBF). A catheter, to permit selective intrarenal administration of BK, was advanced into the proximal left renal artery. The femoral artery was cannulated to measure mean arterial pressure (MAP). MAP, heart rate (HR), and RBF were recorded from conscious unrestrained rats while five-point cumulative dose-response curves during an intrarenal infusion of BK (5-80 microg x kg(-1) x min(-1)) were constructed. Intrarenal infusion of BK elicited dose-dependent increases in MAP (maximum pressor response, 26+/-3 mmHg), accompanied by a significant tachycardia (130+/-18 beats/min) and a 28% increase in RBF. Ganglionic blockade abolished the BK-induced increases in MAP (maximum response, -6+/-5 mmHg), HR (maximum response 31+/-14 beats/min), and RBF (maximum response, 7+/-2%). Selective intrarenal B2-receptor blockade with HOE-140 (50 microg/kg intrarenal bolus) abolished the increases in MAP and HR observed during intrarenal infusion of BK (maximum MAP response, -2+/-3 mmHg; maximum HR response, 15+/-11 beats/min). Similarly, the increases in RBF were prevented after HOE-140 treatment. In fact, after HOE-140, intrarenal BK produced a significant decrease in RBF (22%) at the highest dose of BK. Results from this study show that the cardiovascular responses elicited by intrarenal BK are mediated predominantly via a B2-receptor mechanism.

Systemic reactions to allergy skin tests.

BACKGROUND: Skin testing is a common diagnostic tool in allergy. It is considered a safe procedure, although systemic reactions have been reported. OBJECTIVE: To identify the systemic reaction rates of allergy skin tests and to determine the clinical outcome of such reactions. METHOD: This retrospective study used a computerized database at the Mayo Clinic to identify patients who developed systemic reactions to skin tests. Altogether 497,656 skin tests were performed on 18,311 patients from January 1992 to June 1997. Skin puncture tests were performed on 16,505 patients. Skin puncture and intradermal skin tests were performed on 1,806 patients. Systemic reactions were evaluated and treated by physicians. RESULTS: There were 6 systemic reactions, an overall rate of 33 systemic reactions per 100,000 skin tests. All six patients had asthma. The systemic reaction rates for latex skin testing was 152 or 228 reactions per 100,000 latex skin tests, to penicillin and antibiotics 72 reactions per 100,000 penicillin and antibiotics skin tests, and to aeroallergens 15 or 23 reactions per 100,000 aeroallergen skin tests. The systemic reaction rate for skin puncture test was 30 reactions per 100,000 skin puncture tests, for skin puncture and intradermal skin tests, the rate was 55 reactions per 100,000 skin puncture and intradermal skin tests. All 6 patients were treated and dismissed within 1 hour after treatment. CONCLUSION: The systemic reaction rate to skin tests was very low. Systemic reactions were mild and all patients recovered fully within 1 hour.

Bradykinin B2-receptors mediate the pressor and renal hemodynamic effects of intravenous bradykinin in conscious rats.

Bradykinin (BK) is a peptide which evokes remarkably different changes in cardiovascular function. Systemic bolus injection of BK results in a rapid drop in blood pressure via an endothelium-dependent mechanism. On the other hand, local administration of BK can activate a powerful pressor reflex by stimulating afferent nerves located in the abdominal viscera, the heart, and the kidney. In the present study, the cardiovascular and renal hemodynamic effects during sustained (intravenous infusion) and transient (intravenous bolus injection) elevations in circulating BK were characterized and the receptor mechanism eliciting these effects was investigated. Mean arterial pressure (MAP), heart rate (HR), and renal blood flow (RBF) were recorded from conscious unrestrained rats while five-point cumulative dose-response curves were constructed during infusion or bolus injection of BK (5-80 microg kg(-1)). Infusion of BK produced dose-dependent increases in MAP (maximum response = 27 +/- 3 mmHg) accompanied by a significant tachycardia (maximum response = 159 +/- 20 bpm), a 28 +/- 6% increase in RBF, and no changes in renal vascular resistance (RVR). The BK-induced increases in MAP, HR, and RBF were abolished after treatment with a ganglion blocker (maximum responses: MAP = 2 +/- 3 mmHg, HR = 13 +/- 4 bpm, RBF = 4 +/- 2%) or with an agent which blocks B2-receptors (maximum responses: MAP = 1 +/- 1 mmHg, HR = 6 +/- 5 bpm, RBF = 6 +/- 2%). In marked contrast, bolus administration of BK resulted in hypotensive responses (maximum decline in MAP = -37 +/- 4 mmHg), reflex tachycardia (maximum increase in HR = 45 +/- 9 bpm), increases in RBF (maximum response = 13 +/- 4%), and significant reductions in RVR (maximum response = 38 +/- 5%). These responses were also prevented when B2-receptors were blocked (maximum responses: MAP = 3 +/- 2 mmHg, HR = 17 +/- 6 bpm, RBF = 3 +/- 3%, RVR = 9 +/- 4%). In summary, BK infusions activated a cardiopressor reflex while BK injections caused hypotension. These opposite effects were both mediated via B2-receptors. These findings suggest that BK can have complex effects on the cardiovascular system that may be dependent on the sites, magnitude, and duration of elevated BK concentrations.

Effect of acute increases in filtered HCO3- on renal hydrogen transporters: II. H(+)-ATPase.

Adaptive increases in renal bicarbonate reabsorption occur in response to acute increases in filtered bicarbonate (FLHCO3). In a previous study, we showed that an increase in FLHCO3 induced by plasma volume expansion increased the Vmax for Na+/H+ exchange activity in renal cortical brush border membrane vesicles (BBMV), providing a potential mechanism for the adaptive increase in HCO3- reabsorption. The present studies were undertaken to determine whether the increase in FLHCO3 induced by plasma expansion also stimulates the other major H+ transporter in cortical BBMV, the H(+)-ATPase. H(+)-ATPase activity was assessed in BBMV obtained from hydropenic and plasma expanded Munich-Wistar rats, using a NADH-linked ATPase assay. H(+)-ATPase activity was measured as the ouabain and oligomycin-insensitive, bafilomycin A1-sensitive component of total ATPase activity. Acute plasma expansion doubled single nephron FLHCO3, and this change was associated with a 64% increase in the Vmax for H(+)-ATPase activity, with no change in apparent Km. The Vmax for H(+)-ATPase activity correlated directly with whole kidney GFR and FLHCO3 (r = 0.68 and 0.72, respectively), and with single nephron GFR and FLHCO3 (r = 0.76 and 0.80, respectively). Thus, the mechanism for the adaptive increase in proximal tubular HCO3- reabsorption that occurs in response to acute increases in FLHCO3 appears to be related to increased activity of both H(+)-ATPase and Na+/H+ exchange in the apical membrane of the proximal tubule epithelium.

Cryofixation, cryosubstitution, and immunoelectron microscopy: potential role in diagnostic pathology.

In immunoelectron microscopic investigations, retention of antigenic sites is crucial. Methods for preparing samples for conventional electron microscopy involve chemical fixatives followed by dehydration in organic solvents and embedding in plastic resins, all procedures potentially detrimental to antigenicity. Cryomethods provide a physical fixation alternative for the preparation of biological samples for ultrastructural, immunocytochemical, and microanalysis studies without the use of any chemicals. This can be particularly useful in diagnostic pathology, providing an alternative to conventional fixation methods which sometimes destroy the antigen in question. The recent development of a portable cryofixation device, the PS1000 Portable Metal Mirror Ultra-Rapid Cryofixation Unit (Delaware Diamond Knives, Inc., Wilmington, DE, USA), provides an opportunity to freeze tissue immediately after procurement for use in diagnostic immunocytochemistry studies. This feasibility study examined the quality of tissue preservation with this device, in terms of both preservation of cellular ultrastructure and immunolabeling. Human tonsil and thymus tissue was slam frozen and, after cryosubstitution in Lowicryl K11M, was examined by immunoelectron microscopy. Good ultrastructural preservation was obtained and reasonable immunolabeling with antibodies to AE1/AE3 keratin filaments was also observed.

Spells: in search of a cause.

OBJECTIVE: To determine the cause of spells, present clinical features, and discuss diagnostic approaches. DESIGN: Relevant medical literature is reviewed, and three illustrative cases are presented. RESULTS: Spells are a sudden onset of a symptom or symptoms that are stereotypic, self-limited, and recurrent. A spell involves both subjective perceptions and objective findings. In the assessment of patients who have spells, use of a systematic approach is important in determining the cause. The causes of spells include endocrine, cardiovascular, psychologic, pharmacologic, neurologic, and other miscellaneous disorders. A comprehensive history, physical examination, and basic laboratory studies are important in the initial assessment. Specialized testing is usually needed and directed by clinical suspicion based on the spell "phenotype" (for example, a pheochromocytoma, carcinoid syndrome, or mast cell disease) and the type of facial flush or pallor. CONCLUSION: In the assessment of the patient who has spells, the clinician should cast a wide but defensible diagnostic net. Initial studies should be directed by the clues obtained from the history and physical examination.

Increased tolerance to two oomycete pathogens in transgenic tobacco expressing pathogenesis-related protein 1a.

Expression of pathogenesis-related protein 1a (PR-1a), a protein of unknown biochemical function, is induced to high levels in tobacco in response to pathogen infection. The induction of PR-1a expression is tightly correlated with the onset of systemic acquired resistance (SAR), a defense response effective against a variety of fungal, viral, and bacterial pathogens. While PR-1a has been postulated to be involved in SAR, and is the most highly expressed of the PR proteins, evidence for its role is lacking. In this report, we demonstrate that constitutive high-level expression of PR-1a in transgenic tobacco results in tolerance to infection by two oomycete pathogens, Peronospora tabacina and Phytophthora parasitica var. nicotianae.

Effect of acute changes in glomerular filtration rate on Na+/H+ exchange in rat renal cortex.

Studies were undertaken in Munich-Wistar rats to assess the influence of changes in filtered bicarbonate (FLHCO3), induced by changes in GFR, on Na+/H+ exchange activity in renal brush border membrane vesicles (BBMV). Whole-kidney and micropuncture measurements of GFR, FLHCO3, and whole-kidney and proximal tubule HCO3 reabsorption (APRHCO3) were coupled with BBMV measurements of H+ gradient-driven 22Na+ uptake in each animal studied. 22Na+ uptake was measured at three Na+ concentration gradients to allow calculation of Vmax and Km for Na+/H+ exchange. GFR was varied by studying animals under conditions of hydropenia, plasma repletion, and acute plasma expansion. The increase in GFR, FLHCO3, and APRHCO3 induced by plasma administration correlated directly with an increase in the Vmax for Na+/H+ exchange in BBMV. The Km for sodium was unaffected. In the plasma-expanded rats, the Vmax for Na+/H+ exchange was 22% greater than in the hydropenic rats (P less than 0.025) whereas APRHCO3 was 86% greater (P less than 0.001). These results indicate that increases in FLHCO3, induced by acute increases in GFR, stimulate Na+/H+ exchange activity in proximal tubular epithelium. This stimulation is a mechanism which can, in part, account for the delivery dependence of proximal bicarbonate reabsorption.

Insecticidal effects of an insect-specific neurotoxin expressed by a recombinant baculovirus.

The scorpion Androctonus australis has a peptide (AaIT) which selectively targets the insect sodium channel. This mode of action is similar to that of many widely used chemical insecticides. When Bombyx mori larvae were infected with a recombinant baculovirus carrying a synthetic AaIT gene, the expressed protein was secreted into the hemolymph and caused symptoms consistent with sodium channel blocking, including tremors and feeding cessation at 40 hr p.i. followed by paralysis and death by 60 hr p.i. Larvae infected with control virus died by 96 hr p.i. These results indicate that foreign genes can be used in recombinant baculoviruses to reduce insect feeding damage and increase the rate of insect kill.

Development of a recombinant baculovirus expressing an insect-selective neurotoxin: potential for pest control.

Recombinant nuclear polyhedrosis viruses (NPVs) expressing insect-selective toxins, hormones, or enzymes could enhance their insecticidal properties. We have constructed a recombinant, polyhedrin-positive Autographa californica NPV (AcNPV) that is orally infectious and expresses an insect-selective toxin (AaIT), isolated from the scorpion Androctonus australis, under the control of the p10 promoter. Bioassays with the recombinant baculovirus on 2nd instar larvae of Heliothis virescens demonstrated a significant decrease in the time to kill (LT50 88.0 hours) compared to wild-type AcNPV (LT50 125 hours). Production of AaIT was confirmed by western blot analysis of larval hemolymph from infected H. virescens, and bioassays with larvae of Sarcophaga falculata.

Delivery dependence of early proximal bicarbonate reabsorption in the rat in respiratory acidosis and alkalosis.

In the intact rat kidney, bicarbonate reabsorption in the early proximal tubule (EP) is strongly dependent on delivery. Independent of delivery, metabolic acidosis stimulates EP bicarbonate reabsorption. In this study, we investigated whether systemic pH changes induced by acute or chronic respiratory acid-base disorders also affect EP HCO3- reabsorption, independent of delivery (FLHCO3, filtered load of bicarbonate). Hypercapnia was induced in rats acutely (1-3 h) and chronically (4-5 d) by increasing inspired PCO2. Hypocapnia was induced acutely (1-3 h) by mechanical hyperventilation, and chronically (4-5 d) using hypoxemia to stimulate ventilation. When compared with normocapneic rats with similar FLHCO3, no stimulation of EP or overall proximal HCO3 reabsorption was found with either acute hypercapnia (PaCO2 = 74 mmHg, pH = 7.23) or chronic hypercapnia (PaCO2 = 84 mmHg, pH = 7.31). Acute hypocapnia (PaCO2 = 29 mmHg, pH = 7.56) did not suppress EP or overall HCO3 reabsorption. Chronic hypocapnia (PaCO2 = 26 mmHg, pH = 7.54) reduced proximal HCO3 reabsorption, but this effect was reversed when FLHCO3 was increased to levels comparable to euvolemic normocapneic rats. Thus, when delivery is accounted for, we could find no additional stimulation of proximal bicarbonate reabsorption in respiratory acidosis and, except at low delivery rates, no reduction in bicarbonate reabsorption in respiratory alkalosis.

Metabolic acidosis stimulates bicarbonate reabsorption in the early proximal tubule.

The early proximal tubule is the major site for renal bicarbonate reabsorption but little is known about the influence of acidosis on transport in this segment. This study examined early proximal bicarbonate reabsorption in rats with chronic metabolic acidosis (MA) (induced by NH4Cl administration). Rats were studied by free-flow micropuncture techniques, after varying degrees of plasma volume expansion to vary the filtered load of bicarbonate (FLHCO3). At FLHCO3 less than 700 pmol/min, both control and acidotic animals reabsorbed greater than 80% of the filtered load by 2 mm from Bowman's space. At higher FLHCO3 (700-1,100 pmol/min), reabsorption in the early proximal tubule was significantly greater in MA rats vs. control (633 +/- 26 vs. 449 +/- 24 pmol/min, between 1 and 2 mm from Bowman's space, P less than 0.001). This MA-induced stimulation of early proximal bicarbonate reabsorption was completely reversed by restoring systemic pH to normal either by acute hypocapnia or alkali infusion. Thus bicarbonate reabsorption in the early proximal tubule correlated closely with changes in systemic pH in rats with MA when bicarbonate delivery was increased by plasma expansion. The mechanism of this effect remains to be determined.

Analysis of PCO2 variations in the renal cortex. I. Single nephron.

A mathematical model was developed to predict differences in CO2 partial pressure between afferent arterioles and peritubular capillaries, based on the flow rate and composition of afferent arteriolar blood. Buffering reactions in blood were described by use of conditions of chemical equilibrium and electroneutrality in separate plasma and red cell compartments, with inclusion of such factors as the effect of hemoglobin oxygenation (alkaline Bohr effect) and formation of carbamino compounds. Steady-state mass balance equations allowed the prediction of peritubular capillary blood composition based on the inputs of blood from the efferent arteriole and the addition of water, CO2, NaHCO3, and NaCl derived from tubule reabsorbate. Models developed previously to describe the rates of glomerular filtration, and of proximal tubule reabsorption of HCO3- and CO2, were combined with the peritubular capillary model to allow realistic simulations for a single superficial nephron. The predicted difference of 5.5 mmHg between the CO2 partial pressures in peritubular capillaries and afferent arterioles (delta PCO2) was in good agreement with values reported for normal Munich-Wistar rats. For a given afferent arteriolar blood composition, the calculated delta PCO2 generally decreased with increasing blood flow rate. At a given blood flow rate and afferent PCO2, delta PCO2 decreased as afferent plasma HCO3- concentration was increased. When afferent PCO2 was varied at constant blood flow rate and HCO3- concentration, delta PCO2 changed in parallel with afferent PCO2.