Conditional expression of interferon-gamma to enhance host responses to pulmonary bacterial infection.

Strategies to augment host defense against pulmonary infection run the risk of inducing excess pulmonary inflammation and tissue injury. To address this problem, we investigated conditional expression in lung tissue of the murine interferon-gamma (IFN-gamma) transgene. A recombinant adenoviral vector (AdTetIFN) was constructed by placing a murine IFN-gamma cDNA downstream of a tetracycline (Tet)-responsive promoter, inserted into a replication-defective adenoviral vector. Co-infection of target cells with AdTetIFN and a second vector encoding a reverse tetracycline controlled transactivator allowed doxycycline (Dox)-regulated IFN-gamma production. We then administered 10(8) plaque-forming units (PFU) of AdTetIFN to mice by intratracheal injection. When the mice were provided with Dox in drinking water (0.5mg/ml in 5% sucrose), there was significant release of IFN-gamma in lavage fluid by ELISA in comparison to mice on water/sucrose alone (399+/-74 pg/ml vs undetectable, p<0.01). IFN-gamma in lavage fluid was associated with upregulation of Class II Major histocompatibility complex markers on alveolar macrophages by flow cytometry, suggesting macrophage activation. We then injected AdTetIFN into mice three days prior to challenge with 10(4) CFU Klebsiella pneumoniae. Test mice were maintained on water+Dox and control mice on water+sucrose. Bacterial burden was assayed in lung tissue at serial intervals. At 24h after challenge, mice on doxycycline had significantly lower infection burden in comparison to mice on water/sucrose (0.77+/-0.05 colony forming units/lung for 10(8) PFU AdTetIFN plus Dox compared to 1.4+/-0.11 colony-forming units/lung for AdTetIFN without Dox, p<0.05). Survival of the vector treated mice given doxycycline in drinking water was also enhanced. Microscopic examination of lavaged cells showed a significant increase in pulmonary neutrophils in the AdTetIFN+Dox mice in comparison to AdTetIFN+sucrose mice (16+/-1.0 x 10(5) vs 10+0.8 cells/lung, p<0.05). We conclude that local release of IFN-gamma can be selectively activated to enhance neutrophil recruitment and host resistance to bacterial pneumonia.

Tight control of transgene expression by lentivirus vectors containing second-generation tetracycline-responsive promoters.

BACKGROUND: The goal of this study was to design improved regulatable lentivirus vector systems. The aim was to design tetracycline (tet)-regulatable lentivirus vectors based on the Tet-on system displaying low background expression in the absence of the doxycycline (DOX) inducer and high transgene expression levels in the presence of DOX. METHODS: We constructed a binary lentivirus vector system that is composed of a self-inactivating (SIN) lentivirus vector bearing inducible first- or second-generation tet-responsive promoter elements (TREs) driving expression of a transgene and a second lentivirus vector encoding a reverse tetracycline-controlled transactivator (rtTA) that activates transgene expression from the TRE in the presence of DOX. RESULTS: We evaluated a number of different rtTAs and found rtTA2S-M2 to induce the highest levels of transgene expression. Regulated transgene expression was stable in human breast carcinoma cells implanted into nude mice for up to 11 weeks. In an attempt to minimize background expression levels, the chicken beta-globin cHS4 insulator element was cloned into the 3' long terminal repeat (LTR) of the transgene transfer vector. The cHS4 insulator element reduced background expression but expression levels following DOX addition were lower than those observed with vectors lacking an insulator sequence. In a second strategy, vectors bearing second-generation TREs harboring repositioned tetracycline operator elements were used. Such vectors displayed greatly reduced leakiness in the absence of DOX and induced transgene expression levels were up to 522-fold above those seen in the absence of DOX. CONCLUSIONS: Inducible lentivirus vectors bearing insulators or second-generation TREs will likely prove useful for applications demanding the lowest levels of background expression.

Sulbactam plus ampicillin: interim review of efficacy and safety for therapeutic and prophylactic use.

The efficacy and safety of sulbactam/ampicillin has been evaluated in 39 studies of therapeutic use and six studies of prophylaxis. Studies of therapy were conducted in 899 patients: 751 seriously ill, many of whom had multiple concurrent diseases, and 148 with gonorrhea. Overall clinical and bacteriologic success was achieved in 92% of assessable cases; 88% of 768 pathogens in these patients were eradicated. Of these pathogens, 43% were resistant to ampicillin; eradication rates of 91% and 85% were achieved in ampicillin-resistant and ampicillin-sensitive organisms, respectively. In 388 patients who received prophylactic sulbactam/ampicillin, efficacy was similar to that of comparative agents and better than that of a placebo in preventing wound infections after appendiceal, biliary, upper-gastrointestinal, or gynecologic surgery. Adverse reactions were infrequent with the exception of injection-site pain, which occurred mainly after intramuscular injection and was reduced in incidence by concurrent administration of lidocaine.

The antihypertensive effects of doxazosin: a clinical overview.

Doxazosin is a long-acting selective alpha 1-adrenoceptor antagonist which has been shown to be effective and well tolerated in the treatment of hypertension given in once-daily doses as monotherapy for up to 1 year or as an adjunct to thiazide or beta-adrenoceptor blockers. Doxazosin has a pharmacokinetic profile in both young adult and elderly subjects which is compatible with once-daily administration. This has been confirmed by measurement of steady state pharmacokinetics in patients receiving long-term doxazosin therapy. In controlled double-blind studies involving approximately 550 patients on doxazosin 1-16 mg once daily, significant reductions in both standing and supine BP were maintained throughout the 24 h dosing interval. Effectiveness of doxazosin in terms of BP lowering and proportion of responders was similar to that achieved with hydrochlorothiazide 25-100 mg once daily, atenolol 50-100 mg once daily, nadolol 40-160 mg once daily, metoprolol 100-200 mg per day given twice daily, or prazosin 1-20 mg per day given twice daily. Doxazosin was as effective in elderly patients as in the younger age group and was as effective in blacks as in caucasians. Doxazosin was well tolerated. Side-effects were generally mild to moderate in severity. Overall incidence, including postural effects early in treatment, was similar to that seen with the comparative agents. In comparison with placebo, doxazosin favourably increased (P less than 0.05) the HDL/total cholesterol ratio.(ABSTRACT TRUNCATED AT 250 WORDS)

Endocrine abnormalities in a patient with partial trisomy 4q.

Partial trisomy of the long arm of chromosome 4, usually resulting from a familial segregation of a balanced translocation, has been described in a number of patients. This report describes the genetic and endocrine findings in a 16 year old 46,XY,12q+ mentally retarded male. The banding pattern of the extra chromatin material from this de novo unbalanced translocation shows that the distal segment of the long arm of chromosome 4 is involved. Comparison of the clinical features in this patient with cases of partial trisomy 4q previously reported support the cytogenetic evidence for this translocation involving the distal portion of 4q. Endocrine data suggested an end-organ resistance, characterised by extreme hyperinsulinaemia, primary hypothyroidism, and hypergonadotrophic hypogonadism associated with no signs of autoimmunity. To our knowledge, no endocrine evaluation has been previously reported in patients with partial trisomy 4q.

A re-evaluation of the mechanisms underlying simple cell orientation selectivity.

Following from evidence supporting GABA as a putative inhibitory transmitter in the visual cortex, we have iontophoretically applied the GABA antagonist N-methyl bicuculline (Nmb) to simple cells in order to block the inhibitory inputs acting on them. We found that under these conditions previously sharply-tuned simple cells responded equally to all orientations. Moreover receptive field dimensions, judged by the response to stimuli at the optimal and orthogonal orientations, equated best with that expected from a single dLGN cell input. It seems thus, that asymmetries in the excitatory input are not a significant factor in the generation of simple cell orientation selectivity. The asymmetry underlying orientation selectivity rather originates from the operation of an intracortical inhibitory mechanism.

Effects of drugs acting on cerebral 5-hydroxytryptamine mechanisms on dopamine-dependent turning behaviour in mice.

1. The effects of drugs acting on cerebral 5-hydroxytryptaminergic mechanisms on drug-induced turning behaviour in mice with unilateral destruction of nigro-striatal dopaminergic nerve terminals have been studied. 2. Administration of L-tryptophan (400 mg/kg) or 5-hydroxytryptophan (200 mg/kg) increased brain 5-hydroxytryptamine and decreased the turning induced by both apomorphine (2 mg/kg) and amphetamine (5 mg/kg). 3. Parachlorophenylalanine (3 X 500 mg/kg) decreased brain 5-hydroxytryptamine and increased both apomorphine and amphetamine-induced circling behaviour. 4. Varying the protein content of dietary intake significantly altered brain 5-hydroxytryptamine and tryptophan levels, spontaneous locomotor activity and amphetamine-induced circling behaviour in these mice. 5. Systemic administration of methysergide (0.5-4 mg/kg), lysergic acid diethylamide (0.025-0.2 mg/kg), cyproheptadine (2.5-20 mg/kg) or clomipramine (0.6-20 mg/kg) produced no consistent effect on drug-induced turning behaviour. 6. The results suggest that circling behaviour due to striatal dopamine receptor stimulation is depressed by an elevation of brain 5-hydroxytryptamine and enhanced by a reduction in brain 5-hydroxytryptamine. 7. The possible physiological relationship between dopamine and 5-hydroxytryptamine neurones in the basal ganglia is discussed.