Live imaging of Toll-like receptor 2 response in cerebral ischaemia reveals a role of olfactory bulb microglia as modulators of inflammation.

Activation of microglial cells in response to ischaemic injury, inflammatory and/or immune stimuli is associated with the marked induction of Toll-like receptor 2 (TLR2). At present, little is known about the spatial and temporal sequence of events, micro-regional specificities and the potential long term role of the TLR2 response to brain injuries. To investigate microglial activation/TLR2 response in real time, we generated a transgenic mouse model bearing the dual reporter system luciferase/green fluorescent protein under transcriptional control of a murine TLR2 promoter. In this model, transcriptional activation of TLR2 was visualized in the brains of live animals using biophotonic/bioluminescence molecular imaging and a high resolution/sensitivity charged coupled device camera. It was found that TLR2 induction/microglial activation has a marked chronic component after ischaemic injury and may last several months after the initial attack. The pro-inflammatory response was not restricted to the site of ischaemic injury but was also evident in the olfactory bulb. A significant TLR2 response was first seen in the olfactory bulb 6 h after stroke and several hours before the increase in photon emission over the site of infarction. This sequence of events was further confirmed by immunohistochemistry. A similar early TLR2 response from olfactory bulb microglia was observed in the brain's immune response to pathogens. We therefore propose that, owing to their unique situation, receiving and translating numerous inputs from the brain as well as from the environment, olfactory bulb microglia may serve as sensors and/or modulators of brain inflammation.

Type 1 hereditary tyrosinemia. Evidence for molecular heterogeneity and identification of a causal mutation in a French Canadian patient.

Type 1 hereditary tyrosinemia (HT1) is a metabolic disorder caused by a deficiency of fumarylacetoacetate hydrolase (FAH). Using a full-length FAH cDNA and specific antibodies, we investigated liver specimens from seven unrelated HT1 patients (six of French Canadian and one of Scandinavian origin). The expression of FAH in livers of these individuals was analyzed at several molecular levels including mRNA, immunoreactive material (IRM), and enzymatic activity. Four phenotypic variants were differentiated by these assays: (i) presence of FAH mRNA without any IRM or enzymatic activity, (ii) decreased FAH mRNA, IRM, and enzymatic activity, (iii) moderately decreased FAH mRNA and IRM with severely reduced enzymatic activity, and (iv) undetectable FAH mRNA, IRM, and enzymatic activity. These various molecular phenotypes suggest that this disorder may be caused by a variety of FAH mutations. Interestingly, we found no apparent relationship between the clinical and the molecular phenotypes, except that patients with absent IRM and enzymatic activity tend to have higher levels of serum alpha-fetoprotein and an earlier clinical onset. To further analyze the molecular basis of HT1, the FAH cDNA of a patient designated as variant A was amplified and sequenced. An A-to-T transversion, which substitutes asparagine16 with isoleucine (N16I), was identified. This patient was heterozygous as shown by direct sequencing of the amplified region and hybridization with allele-specific oligonucleotide probes. The N16I allele originates from the father and the second allele appears not to be expressed in the liver of the proband. CV-1 cells transfected with the mutant cDNA produced FAH mRNA, but no protein or hydrolytic activity, as predicted by the "A" phenotype of the patient. This is the first demonstration of heterogeneity in the expression of FAH at the levels of protein, mRNA, and enzymatic activity in the livers of HT1 patients and is the first identification of a causal mutationin this disease.

Thallium-201 myocardial imaging during pharmacologic coronary vasodilation: comparison of oral and intravenous administration of dipyridamole.

Although the diagnostic utility of thallium-201 myocardial imaging after dipyridamole infusion is well established, the intravenous form of the drug is not yet commercially available in North America. Fifty patients referred for coronary angiography were prospectively studied. Within a 2 week period, each patient underwent cardiac catheterization and thallium-201 myocardial imaging after both oral and intravenous dipyridamole administration. For the oral protocol, patients were randomly assigned to treatment with either 200 or 400 mg of dipyridamole in tablet form. Coronary artery stenoses of 70% or greater were considered significant. For the 25 patients who received a 200 mg oral dose of dipyridamole, the scintigraphic study showed perfusion defects in 65% of patients with significant coronary artery disease after the oral dose and in 85% of patients after the intravenous dose. For the 25 patients who received a 400 mg oral dose, the sensitivity of the scintigram was 84% after the oral dose and 79% after the intravenous dose. Except for headache and nausea, side effects were less severe and less frequent with oral (either 200 or 400 mg) than with intravenous dipyridamole. Because of the delayed and variable absorption of dipyridamole tablets, the oral studies required a longer period of medical supervision (45 to 60 minutes), and aminophylline was empirically administered after completion of the first set of thallium-201 images. It is concluded from this study that thallium-201 myocardial imaging after coronary vasodilation with a 400 mg oral dose of dipyridamole is a safe, widely available and reliable alternative for the evaluation of coronary artery disease in patients unable to achieve an adequate exercise level on stress testing.

Enalapril versus digoxin in patients with congestive heart failure: a multicenter study. Canadian Enalapril Versus Digoxin Study Group.

Patients with New York Heart Association functional class II or III heart failure stabilized on furosemide therapy were entered into a randomized controlled trial comparing enalapril (n = 72) and digoxin (n = 73). End points were clinical outcome, treadmill exercise capacity and echocardiographic left ventricular dimensions. Improvement in clinical outcome was defined as a reduction of at least one functional class or withdrawal because of an adverse clinical event. After 4 weeks, 13 patients receiving enalapril showed improvement, 55 had no change and 9 manifested deterioration compared with 7, 49 and 17, respectively, in the digoxin group (p less than 0.01). After 14 weeks, 13 patients receiving enalapril showed improvement, 50 had no change and 9 manifested deterioration, compared with 14, 37 and 22, respectively, in the digoxin group (p less than 0.025). More patients in the digoxin group were withdrawn because of an adverse clinical event (p less than 0.05). Exercise time and percent fractional shortening improved in both groups (p less than 0.001 and less than 0.05, respectively), with no significant difference between groups (p greater than 0.50). Both rate-pressure product and subjectively evaluated exertion during submaximal exercise were reduced only in the enalapril group. Although the majority of patients in both groups did well, those receiving enalapril experienced fewer adverse clinical events and had less fatigue during submaximal exercise.

Relative importance of psychologic traits and severity of ischemia in causing angina during treadmill exercise. Canadian Amlodipine/Atenolol in Silent Ischemia Study (CASIS) Investigators.

OBJECTIVES: This study was conducted to compare the influence of psychologic traits versus ischemia severity on the occurrence of angina during treadmill exercise. BACKGROUND: Some studies suggest that angina is associated with certain psychologic traits, whereas others show an association with more severe ischemia. The relative influence of these two factors and the extent to which they interact are not known. METHODS: Off-drug treadmill exercise testing and a battery of psychologic tests were performed on 122 patients with known coronary artery disease. Psychologic tests measured sensitivity to physical symptoms, denial and deception, type A behavior, anger, hostility, depression, marital adjustment and amount of external stress. Stepwise logistic regression was used to determine the independent association of psychologic traits, ischemic threshold and exercise tolerance with the occurrence of angina. RESULTS: Angina during treadmill exercise was reported by 66 of 122 patients. On univariate testing, angina was positively associated with sensitivity to physical symptoms (p < 0.001), type A behavior (p = 0.021) and depression (p = 0.032) and was negatively associated with exercise tolerance (p < 0.001) and work load threshold for ischemia (p < 0.01). Multivariate analysis revealed independent and additive associations of angina with sensitivity to physical symptoms (p = 0.003), exercise capacity (p = 0.003) and work load threshold for ischemia (p = 0.018). Once these were included in a logistic model, depression and type A behavior were no longer significant. Other psychologic traits showed no association with angina. CONCLUSIONS: Sensitivity to physical symptoms, ischemic threshold and exercise tolerance are independently associated with angina, with sensitivity to physical symptoms having the stronger influence. The physiologic and psychologic mechanisms underlying symptom perception have an influence on angina that is independent of and additive to the severity of underlying ischemia.

Effect of amlodipine, atenolol and their combination on myocardial ischemia during treadmill exercise and ambulatory monitoring. Canadian Amlodipine/Atenolol in Silent Ischemia Study (CASIS) Investigators.

OBJECTIVES: This study compared the effects of amlodipine, atenolol and their combination on ischemia during treadmill testing and 48-h ambulatory monitoring. BACKGROUND: It is not known whether anti-ischemic drugs exert similar effects on ischemia during ambulatory monitoring and exercise treadmill testing. METHODS: Patients with stable coronary artery disease and ischemia during treadmill testing and ambulatory monitoring were randomized to receive amlodipine (n = 51) or atenolol (n = 49). Each group underwent a counterbalanced, crossover evaluation of single drug and placebo, followed by evaluation of the combination. RESULTS: Amlodipine and the combination prolonged exercise time to 0.1-mV ST segment depression by 29% and 34%, respectively (p < 0.001) versus 3% for atenolol (p = NS). During ambulatory monitoring, the frequency of ischemic episodes decreased by 28% with amlodipine (p = 0.083 [NS]), by 57% with atenolol (p < 0.001) and by 72% with the combination (p < 0.05 vs. both single drugs; p < 0.001 vs. placebo). Suppression of ischemia during exercise testing and ambulatory monitoring was similar in patients with and without exercise-induced angina. Exercise time to angina improved by 29% with amlodipine (p < 0.01), by 16% with atenolol (p < 0.05) and by 39% with the combination (p < 0.005 vs. placebo, atenolol and amlodipine). In patients with angina, total exercise time improved by 16% with amlodipine (p < 0.001), by 4% with atenolol (p = NS) and by 19% with the combination (p < 0.05 vs. placebo and either single drug). In those patients without angina, no therapy significantly improved total exercise time. CONCLUSIONS: Ischemia during treadmill testing was more effectively suppressed by amlodipine, whereas ischemia during ambulatory monitoring was more effectively suppressed by atenolol. The combination was more effective than either single drug in both settings.

Effect of aminoglycosides on the disposition of thyroid hormones and thyroglobulin.

Our study was designed to confirm the potential effects of three aminoglycosides on the disposition of thyroid hormones. Twenty-seven patients diagnosed with either cellulitis (n = 19), chronic osteitis (n = 4), or an abscess (n = 4) were selected. Thirteen patients received tobramycin, 60 to 100 mg iv q. 8 h., plus cloxacillin, 1 gm iv q. 4 h.; seven patients received netilmicin, 40 to 120 mg iv q. 8 h., plus cloxacillin, 1 gm iv q. 4 h.; and seven patients received either cloxacillin, 1.5 gm iv q. 4 h., or cefoperazone, 2 to 4 gm iv q. 12 h. for at least 7 days. Another group of six normal subjects received neomycin, 0.5 gm po q. 6 h. for 7 days. All these subjects had normal thyroid function before antibiotic dosing and none had thyroid function abnormalities. Tobramycin and cloxacillin/cefoperazone did not influence thyroid function. Netilmicin decreased the total serum concentrations of triiodothyronine (T3) from 114 +/- 9 to 75 +/- 7 ng/dl (P less than 0.01), probably because of increased clearance, as the T3 free fraction increased from 0.43% +/- 0.02% to 0.49% +/- 0.02% (P less than 0.05). Thyroxine (T4) and reverse T3 (rT3) levels were not affected. Neomycin decreased T3 levels from 104 +/- 8 to 92 +/- 7 ng/dl (P less than 0.05) and the serum concentrations of thyroglobulin from 17.3 +/- 2.0 to 11.7 +/- 2.0 ng/ml (P less than 0.001). Because T4 and rT3 levels did not change, our results suggest that neomycin may have directly affected the gland. We conclude that some aminoglycosides can alter the disposition of thyroid hormones.

Cloning and expression analysis of a cDNA encoding fumarylacetoacetate hydrolase: post-transcriptional modulation in rat liver and kidney.

Fumarylacetoacetate hydrolase (FAH) is an enzyme which is deficient in human hereditary tyrosinemia type 1. We have cloned and sequenced a rat liver cDNA encoding FAH. The identity of the clone was ascertained by hybrid-selection experiments and deduced amino acid (aa) sequence homologies with sequenced oligopeptide fragments of the purified rat liver protein. The cDNA codes for a 419-aa protein of 45,946 daltons. We used this cDNA as a probe in conjunction with a specific anti-rat FAH antibody to study the expression pattern of the FAH gene in rat liver and kidney. Northern blot analysis indicates that the kidney contains slightly more FAH mRNA that the liver. Western blotting shows, however, that the liver contains about twice as much FAH protein as the kidney. Primer extension experiments suggest that there are no differences in the 5'-untranslated (UT) ends of the FAH mRNA of both tissues. We conclude that synthesis of the FAH protein is in part regulated at the post-transcriptional level in rats liver and kidney, and that this regulation does not appear to be mediated by the 5'-UT sequence of the FAH mRNA.

Comparison between thallium-201, technetium-99m-sestamibi and technetium-99m-teboroxime planar myocardial perfusion imaging in detection of coronary artery disease.

Technetium-99m-sestamibi (MIBI) and 99mTc-teboroxime (TEBO) are two new myocardial perfusion imaging agents. The purpose of this prospective study was to compare MIBI and TEBO to 201TI planar imaging. Eighteen patients with significant coronary artery disease on coronary angiogram were submitted to three treadmill stress tests performed within 3 mo and were imaged with the three radiopharmaceuticals as follows. 1. TI: 2.2 mCi, immediate and delayed views (4 hr later, 8 min/view). 2. TEBO: 15-20 mCi at stress (1 min/view) and a second injection was repeated 4 hr later at rest (20-25 mCi). 3. MIBI: 15-18 mCi at stress (8 min/view) and 1-4 days later, 15-18 mCi at rest. Patients achieved similar levels of exercise. A blinded reading was performed by three observers. The left ventricle was divided into three segments/view and ischemic/normal wall ratios were also determined. Segmental comparison showed an agreement in 85% (138/162) of the segments between TI and TEBO, in 92% (149/162) between TI and MIBI and in 84% (136/162) between MIBI and TEBO. Abnormal TI, MIBI and TEBO studies were seen in 16 (89%), 16 (89%) and 15 (83%) patients, respectively, detecting 77, 75 and 65 abnormal segments. Ischemic-to-normal wall ratios were 0.75 +/- 0.06, 0.73 +/- 0.08 and 0.78 +/- 0.08 for TI, MIBI and TEBO, respectively. In conclusion, although the biologic characteristics of these agents are different, this study showed a good correlation between them in detection of significant coronary artery disease (high pretest likelihood population).

Sources of nitrate exposure in residents of rural areas in Quebec, Canada.

Nitrate exposure was investigated in a group of 187 people using well water and living in four areas of rural Quebec (Canada) with intensive agricultural activities. Nitrate intake was evaluated using a 24-h dietary recall and a food frequency questionnaire, in conjunction with a validated food database and measurements of nitrate concentrations in private wells. The total internal dose was estimated by means of the 24-h urinary nitrate excretion, while taking into account risk factors for endogenous nitrate formation. Mean (geometric) 24-h urinary nitrate excretion was 16.9 mg N for the 100 people with low groundwater contamination (mean nitrate concentration=0.18 mg N/l) and 23.3 mg N in the 87 individuals with moderate groundwater contamination (mean nitrate concentration=7.1 mg N/l). A multivariate analysis revealed that dietary nitrate intake during the last 24 h was the principal source of exposure, followed by water intake during the last 24 h. The Quetelet index was also a significant predictor of urinary excretion. The total predictive model explained only 29% of the variability in urinary nitrate excretion (R2=0.286). Neither the inflammatory status as indicated by elevated C reactive protein, the presence of Helicobacter pylori antibodies nor the occurrence of diarrhea during the last 24 h prior to urine collection were associated with urinary nitrate excretion. In conclusion, food and to a lesser extent water contribute to nitrate exposure in this rural setting with moderate water contamination. Better predictors of endogenous nitrate production are needed to improve our ability to model nitrate body burden and estimate associated health risks.

[Contamination of drinking water by nitrates: analysis of health risks]. / La contamination de l'eau potable par les nitrates: analyse des risques à la santé.

Nitrate ground water contamination is a relatively frequent problem due to the massive use of fertilizers in agriculture. The health effects of contamination are due to the transformation of nitrates into nitrites in the G.I. tract and possibly the transformation of nitrites into nitrosamines in the stomach. The risk of methemoglobinemia in infants is due to nitrites contained in the water used to reconstitute milk for feeding. There are possible but unproven risks associated with the formation of nitrosamines, i.e. teratogenous and cancer risks. Drinking water standards are only based on the risk of methemoglobinemia, but given the present state of knowledge they appear to be safer in relation to the other potential risks. In the face of the increase in nitrates in ground water and the partially known risks, it appears necessary to try and prevent this type of contamination and inform the persons at risk, such as pregnant women and young mothers. It would also be useful to promote research on the effects of nitrates and nitrites on humans.

Estrogen receptor alpha-mediated adiposity-lowering and hypocholesterolemic actions of the selective estrogen receptor modulator acolbifene.

OBJECTIVE: The selective estrogen receptor (ER) modulator (SERM) acolbifene (ACOL), a potent and pure antiestrogen in the mammary gland and uterus, exerts beneficial pro-estrogenic actions on energy balance, insulin sensitivity and lipid metabolism. ACOL binds ERs alpha and beta, both of which have been involved in the metabolic actions of estrogen. This study aimed at determining the identity of the ER involved in the beneficial metabolic actions of ACOL. DESIGN AND MEASUREMENTS: ACOL was administered for 4 weeks to male and female wild-type and ERalpha knockout (KO) mice, and indices of energy balance as well as plasma and liver lipid concentrations were determined. RESULTS: ERalpha KO mice were heavier, gained more fat mass and had larger adipose depots than their wild-type counterparts. In both genders, ACOL decreased fat gain (50%) and white adipose tissue mass in male and female wild-type, but not in ERalpha KO mice. ACOL reduced plasma cholesterol in female wild-type mice (-27%), whereas the compound remained ineffective in their ERalpha KO counterparts. Plasma triglycerides were unaffected by ACOL. Finally, ACOL decreased liver cholesterol and triglyceride concentrations only in wild-type female animals. CONCLUSION: The beneficial metabolic actions of the SERM ACOL on adiposity and on plasma and liver lipids are entirely due to its interaction with the ERalpha.

Agar disk diffusion susceptibility characteristics of azlocillin, carbenicillin, mezlocillin, piperacillin, and ticarcillin.

The agar disk diffusion susceptibility of Enterobacteriaceae to mezlocillin and piperacillin was correlated with agar minimal inhibitory concentrations and compared with the susceptibility to carbenicillin. The agar disk susceptibility of Pseudomonas aeruginosa to azlocillin, mezlocillin, and piperacillin was correlated with agar minimal inhibitory concentrations and compared with the susceptibility to carbenicillin and ticarcillin. Criteria are offered for the zones of inhibition to provide information about resistant and susceptible isolates that correlate with known serum levels.

In vitro antagonism between N-formimidoyl thienamycin and aztreonam, ticarcillin and ticarcillin/clavulanic acid.

One-hundred eighty-seven bacterial strains were tested by the two-disk-agar diffusion method for the interaction between N-formimidoyl thienamycin and aztreonam, ticarcillin and ticarcillin-clavulanic acid. An antagonism between N-formimidoyl thienamycin and the other 3 beta-lactams was noted in half of the evaluable tests, especially against Pseudomonas, Escherichia coli and Morganella.

Contamination of local wildlife following a fire at a polychlorinated biphenyls warehouse in St Basile le Grand, Quebec, Canada.

This study on wildlife contamination, one to ten months after the polychlorinated biphenyls (PCB) fire in St Basile le Grand, Quebec, shows that the fire increased PCB and polychlorinated dibenzofuran (PCDF) levels in animals. From the data, it was not possible to detect a significant increase in polychlorinated dibenzodioxin (PCDD) levels after the fire. Given the relatively small sample size, the differences in concentrations could not be estimated precisely. However, it can be asserted with a 95% confidence level that mean concentrations of total PCBs were roughly 2 to 6 times higher in the area contaminated by the plume of smoke, concentrations of homologues with 3 chlorine atoms were 1 to 4 times higher, and levels of homologues with 5 to 9 chlorine atoms were 3 to 13 times higher. The relative deviations between concentrations in areas under the plume and those outside it were similar for all animals sampled. With regard to total PCDFs, mean concentrations were significantly higher under the smoke plume than outside it for all species. This observation is linked to homologues with 4, 5 and 7 chlorine atoms for which significant differences were detected between the two areas. The fire had no effect on the pattern of PCB congeners found in the tissue of animals in the region. Congeners Nos. 153, 180, 138 and 118 represent approximately 50% of total PCBs. Although PCB and PCDF concentrations were higher in the tissue of local wildlife species exposed to the fire, they were nonetheless comparable to those found in other urban and agricultural areas in Canada. These concentrations, in 2,3,7,8-TCDD toxic equivalents, were much lower than those observed in the wake of three other major incidents involving PCDDs (Elgin, Florida; Times Beach, Missouri; and Seveso, Italy).(ABSTRACT TRUNCATED AT 250 WORDS)

Differential effect of the immunomodulatory hormone somatostatin on replication of human immunodeficiency virus type 1 in CD4+ and CD8+ T lymphocytes.

The long-acting somatostatin analog octreotide (SMS 201-995) possesses immunosuppressive properties and has been successfully used for the management of human immunodeficiency virus (HIV)-associated diarrhea, a condition commonly observed in the absence of known enteric pathogens. Since HIV type 1 (HIV-1) replication can occur in both CD4+ and CD8+ lymphocytes, we hypothesized that this benefit might be due to local effects on HIV-1 replication in these two T-cell subsets. As a model, we studied the effects of two synthetic molecules, SRIH 1-14 and SRIH 1-28, closely related to naturally occurring forms of somatostatin, as well as SMS 201-995 on HIV-1 replication in CD4+ and CD8+ cells derived from peripheral blood mononuclear cells (PBMC). We found that HIV-1 replication was inhibited in CD8+ cells but enhanced in infected CD4+ lymphocytes, as measured by p24 antigen levels in culture fluids. These differential effects were drug concentration dependent. We also observed that somatostatin inhibited the mitogen-induced proliferative responsiveness of both cell types. These effects on both HIV-1 replication and cell proliferation were independent of somatostatin gene expression, since somatostatin mRNAs were not detected in mitogen-stimulated PBMC, as determined by reverse transcription-PCR.

Aztreonam: correlation between disk diffusion and agar plate dilutions susceptibility tests.

Regression lines and error rate-bounded analyses were performed for correlating inhibitory zone diameters with the 30-micrograms aztreonam disk and aztreonam minimal inhibitory concentration for gram-negative aerobic clinical isolates. A correlation coefficient of -0.81 was found for the regression line. Preliminary proposed criteria to distinguish susceptible isolates from resistant ones are: susceptible greater than or equal to 22 mm (MIC less than or equal to 8 micrograms/ml), intermediate 16-21 mm (MIC = 16 micrograms/ml) and resistant less than or equal to 15 mm (MIC greater than or equal to 32 micrograms/ml). Using these criteria, the rates for false-susceptible and false-resistant isolates were zero.

Intravenous injection of an adenovirus encoding hepatocyte growth factor results in liver growth and has a protective effect against apoptosis.

BACKGROUND: Hepatocyte growth factor/scatter factor (HGF/SF) is a pleiotropic cytokine with mitogenic, motogenic and morphogenic effects for a wide variety of cells. Previous studies have reported that the in vivo infusion in normal, untreated mice of recombinant HGF results in low levels of DNA synthesis and liver proliferation. In this study, we examined whether liver regeneration could be obtained by the in vivo injection of a recombinant adenoviral vector encoding human HGF (Ad.CMV.rhHGF) in normal, intact mice. MATERIALS AND METHODS: C57BL/6 mice were infused intravenously with doses increasing from 1 to 4 x 1011 particles of the recombinant human HGF (rhHGF) adenoviral vector or with a control virus encoding Escherichia coli beta-galactosidase (Ad.CMV.lacZ). At day 5, mice were sacrificed and evaluated for the presence of hepatocyte mitogenesis and liver regeneration (5-bromo-2'-deoxyuridine (BrdU) assays and liver weight determination) and for the presence of liver damage (serum alanine amino-transferase (ALT) measurements and TUNEL assays). RESULTS: In vivo administration of rhHGF stimulated DNA synthesis of hepatocytes and liver weight in a dose-dependent fashion. The maximal effect was seen after the infusion of 3 x 1011 particles which resulted at day 5 in >130% increase in relative liver mass with little cytopathic effect. In contrast, administration of the lacZ adenoviral vector caused little hepatocyte replication, but induced high levels of serum ALT (approximately 3 times higher than the rhHGF vector) and significant apoptotic cell death. CONCLUSIONS: This study shows that a single injection of Ad.CMV.rhHGF alone is able to induce in vivo and in a very short period of time, robust DNA synthesis and liver proliferation in normal mice without liver injury or partial hepatectomy. This recombinant adenoviral vector has a lower toxicity than the control lacZ adenovirus. This suggests that HGF may have a protective effect against adenovirus-induced pathology.

Characterization of the human fumarylacetoacetate hydrolase gene and identification of a missense mutation abolishing enzymatic activity.

Hereditary tyrosinemia type 1 is an autosomal recessive disease caused by a deficiency of the last enzyme in the catabolic pathway of tyrosine, fumarylacetoacetate hydrolase (FAH). To analyze the mutations involved in this disease, and as a first step towards elucidating the mechanisms regulating the transcription of the FAH gene, we have isolated and characterized the human gene coding for FAH. The gene contains 14 exons and spans approximately 35 kilobases of DNA. The 5' end of the gene is highly GC-rich, and eleven putative binding sites for the transcription factor Sp 1 were identified in the proximal region of the promoter. We investigated the molecular basis of FAH deficiency in a hereditary tyrosinemia type 1 patient whose liver FAH showed a very low enzymatic activity. Sequencing of the liver FAH cDNA of the patient revealed a C to A transversion in the FAH mRNA, which predicted the replacement of an alanine (A) residue with an aspartic acid (D) residue at position 134 (A134D) of the amino acid sequence of the corresponding protein. Direct sequencing of genomic DNA indicated that the patient was heterozygous for the A134D mutation. The allele that does not carry the A134D mutation was expressed at a very low level in the liver of the patient. Expression of the mutant allele in CV-1 cells confirmed that the A134D mutation was responsible for the lack of enzymatic activity in the liver of the patient.