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P Leone

Publications and source records attributed to P Leone.

At least 55 records · Page 3Linked to original sources
PubMed1998

Peroral gene therapy of lactose intolerance using an adeno-associated virus vector.

Gene therapy is usually reserved for severe and medically refractory disorders because of the toxicity, potential long-term risks and invasiveness of most gene transfer protocols. Here we show that an orally administered adeno-associated viral vector leads to persistent expression of a beta-galactosidase transgene in both gut epithelial and lamina propria cells, and that this approach results in long-term phenotypic recovery in an animal model of lactose intolerance. A gene 'pill' associated with highly efficient and stable gene expression might be a practical and cost-effective strategy for even relatively mild disorders, such as lactase deficiency.

Administration, Oral↗
PubMed1998

In vivo expression of therapeutic human genes for dopamine production in the caudates of MPTP-treated monkeys using an AAV vector.

An adeno-associated virus (AAV) vector, expressing genes for human tyrosine hydroxylase (TH) and aromatic amino acid decarboxylase (AADC), demonstrated significantly increased production of dopamine in 293 (human embryonic kidney) cells. This bicistronic vector was used to transduce striatal cells of six asymptomatic but dopamine-depleted monkeys which had been treated with the neurotoxin MPTP. Striatal cells were immunoreactive for the vector-encoded TH after stereotactic injection for periods up to 134 days, with biochemical effects consistent with dopamine biosynthetic enzyme expression. A subsequent experiment was carried out in six more severely depleted and parkinsonian monkeys. Several TH/aadc-treated monkeys showed elevated levels of dopamine near injection tracts after 2.5 months. Two monkeys that received a beta-galactosidase expressing vector showed no change in striatal dopamine. Behavioral changes could not be statistically related to the vector treatment groups. Toxicity was limited to transient fever in several animals and severe hyperactivity in one animal in the first days after injection with no associated histological evidence of inflammation. This study shows the successful transfection of primate neurons over a period up to 2.5 months with suggestive evidence of biochemical phenotypic effects and without significant toxicity. While supporting the idea of an in vivo gene therapy for Parkinson's disease, more consistent and longer lasting biochemical and behavioral effects will be necessary to establish the feasibility of this appraoch in a primate model of parkinsonism.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗
PubMed1997

Targets for gene therapy of Parkinson's disease: growth factors, signal transduction, and promoters.

Parkinson's disease gene therapy is in its infancy. All studies to date have been in experimental animals and there are no clinical protocols currently approved. Several non-human primate studies however, have been completed and preliminary data appear promising. When dealing with a complex acquired disorder of unknown etiology, gene therapy is likely to provide symptomatic and palliative relief at best and will not be curative. However, if the gene therapy approach has advantages in terms of the risk/benefit ratio, cost and efficacy over current treatments, then it should be brought to clinical trial. This article discusses some future directions and areas of intense investigation at present. The advances in the field over the past five years have been tremendous and it appears possible that before the year 2000, clinical gene therapy trials in Parkinson's disease will be ongoing.

Animals↗
PubMed1997

Adenovirus vector-mediated gene transfer into human epileptogenic brain slices: prospects for gene therapy in epilepsy.

As a first step in the development of a gene therapy approach to epilepsy, we evaluated the ability of adenovirus vectors to direct the transfer into and expression of a marker gene in human brain slices obtained from patients undergoing surgery for medically intractable epilepsy. Following injection of adenovirus vectors containing the Escherichia coli lacZ gene into hippocampal and cortical brain slices, lacZ mRNA, beta-galactosidase protein, and enzymatic activity were detected, confirming successful gene transfer, transcription, and translation into a functional protein. Transfected cells were predominantly glial, with some neurons expressing beta-galactosidase as well. These results support the potential of adenovirus vectors to transfer genetic information into human epileptogenic brain, resulting in expression of the gene into a functional protein. These findings also have implications for the development of gene therapy approaches to certain seizure disorders. A number of potential therapeutic approaches are discussed, including the elevation of inhibitory neurotransmitter or neuropeptide levels, expression or modulation of postsynaptic receptors, and manipulation of signal transduction systems.

Adenoviruses, Human↗
PubMed1997

Long-term evaluation of cellular immunity during antiretroviral therapy and immunization with human immunodeficiency virus type 1 (HIV-1) Env glycoprotein in HIV-1-infected persons.

Cellular immune responses to human immunodeficiency virus type 1 (HIV-1) antigens, microbial recall antigens, and CD3 monoclonal antibody were studied in HIV-1-infected asymptomatic patients in a phase II, double-blind trial of immunization with recombinant HIV-1 gp160 in or not in association with zidovudine. A vigorous and persistent lymphoproliferative response (LPR) to HIV-1 Env antigens was observed in vaccinated patients. Neither Env-specific lymphocyte cytotoxicity nor LPR to recall antigens was significantly influenced by gp160 administration. The induction of LPRs to HIV-1 envelope proteins did not show positive effects on the course of HIV-1 infection. Patients treated with zidovudine alone or in combination with the immunogen showed improvement of T lymphocyte responses and transient reduction of viremia. These results suggest that antiretroviral therapy is more beneficial than immunization with gp160 and should always be considered in association with future vaccination and immunotherapeutic interventions in HIV-1-infected subjects.

Adolescent↗
PubMed1997

Delayed presentation to clinics for sexually transmitted diseases by symptomatic patients. A potential contributor to continuing STD morbidity.

OBJECTIVE: To determine the proportion of symptomatic patients attending public sexually transmitted disease (STD) clinics who fail to seek care within 7 days of the onset of STD symptoms and their self-reported reasons for delays in obtaining care. DESIGN: Interviewers administered 23-item questionnaire to a cross section of STD clinic clients between April and September 1995. SETTING: Five urban STD clinics in the United States (Birmingham, AL; St Paul, MN; San Francisco CA; Seattle, WA; and Raleigh, NC). RESULTS: Of 1,621 patients with genitourinary symptoms, over one third (35% of men, 37% of women) presented to STD clinics only after 1 week or more of symptoms. Men with genital warts (73%) or with nongonococcal urethritis (23.1%) or symptomatic men who were recent contacts to sex partners with STDs were significantly more likely to delay clinic attendance more than a week than men with gonorrhea (6.5%, p < 0.001 for each). Overall 43.8% of women receiving specific clinical diagnoses other than genital warts delayed clinic attendance for more than 1 week. When asked why they delayed clinic attendance, respondents were most likely to respond that they had hoped their symptoms would "go away" (48.5% of men and 49.4% of women who waited a week or more before seeking care). CONCLUSION: A substantial proportion of patients with genitourinary symptoms attending public STD clinics delay seeking care for a week or more, increasing their likelihood of complications and of transmission of infection to others. Interventions to promote more timely clinic attendance may help reduce STD morbidity in the United States.

Adolescent↗
PubMed1997

Patients attending STD clinics in an evolving health care environment. Demographics, insurance coverage, preferences for STD services, and STD morbidity.

OBJECTIVES: To characterize client demographics, sexually transmitted diseases (STD) morbidity, insurance status, reasons for attending public STD clinics, and future preferences for source of STD services. DESIGN: Cross-sectional study of 2,490 clients attending five urban STD clinics for new problems with interviewer-administered 23-item questionnaire and chart review to obtain clinical and laboratory STD diagnoses. RESULTS: Participants were young (51% < 25 years of age), minority (64% nonwhite), poor (43% < or = $10,000/year), and largely uninsured (59% uninsured, 27% private insurance, and 14% Medicaid). Half had previously visited the STD clinic, and 81% had used other providers for non-STD services in the prior 3 years. STD symptoms were cited as the reason for the visit by 63%. The most common factors associated with seeking care at these STD clinics were walk-in services, costs, and confidentiality concerns. STD morbidity was high; 66% of clients were diagnosed with one or more STD. Most (68%) clients preferred to be treated at the STD clinic in the future if they could go anywhere for STD services. CONCLUSIONS: STD clinics see young, minority, poor, and uninsured clients with high STD rates. Even with unlimited future choice, two thirds of the clients surveyed would still prefer to be treated at STD clinics. This study indicates the continuing need for publicly funded, categorical STD clinics in urban areas with high STD morbidity and the importance of easily accessible, confidential, expert STD services from the private sector and managed care organizations.

Adult↗
PubMed1997

Direct gene transfer into human epileptogenic hippocampal tissue with an adeno-associated virus vector: implications for a gene therapy approach to epilepsy.

PURPOSE: Virus vectors capable of transferring genetic information into human cells provide hope for improved therapy in several neurological diseases, including epilepsy. We evaluated the ability of an adeno-associated virus (AAV) vector to transfer and cause expression of a lacZ marker gene in brain slices obtained from patients undergoing temporal lobectomy for control of medically intractable seizures. METHODS: Human brain slices were injected with an AAV vector (AAVlacZ) encoding Escherichia coli beta-galactosidase and incubated for as long as 24 h. The presence of lacZ mRNA. beta-galactosidase protein and enzymatic activity were assayed by reverse transcriptase polymerase chain reaction (rtPCR), immunocytochemistry, and the X-Gal technique, respectively. RESULTS: AAVlacZ directed the expression in human epileptogenic brain of E. coli beta-galactosidase that had functional activity. Expression was observed in < or =5 h and was sustained for as long as the slices were viable. Morphological analysis indicated that neurons were preferentially transfected, and there was no evidence of cytotoxicity. CONCLUSIONS: Our results confirm the feasibility of using AAV vectors to transfer genes into the human CNS and in particular, into neurons. Replacement of the lacZ gene with a functional gene modulating hippocampal neuronal physiology, might allow a localized genetic intervention for focal seizures based on the stereotaxic or endovascular delivery of such a vector system into the appropriate brain region.

Dependovirus↗
PubMed1997

The effect of leptin is enhanced by microinjection into the ventromedial hypothalamus.

To determine whether changes in food intake produced by leptin involve targeting the hormone to distinct central nervous system regions, guide cannulas were positioned stereotaxically into three brain regions--the ventromedial hypothalamus (VMH) (bilaterally, n = 6), the dorsal raphe nucleus (n = 3), and the lateral ventricle (n = 3)--of nonobese male rats (400-500 g). Daily food intake and body weight changes were measured during twice-daily injections of saline (0.1 microl) followed by recombinant human leptin (0.05 microg) for 3 days via the brain cannulas. VMH-injected rats also were followed during a postleptin saline recovery interval. This small dose of leptin did not change food intake or body weight from that during the preceding saline injection period in ventricle-injected or dorsal raphe-injected rats. In sharp contrast, VMH-injected rats ate much less food (56 +/- 8% basal) and lost 9 +/- 3 g/day or 5% of their body weight during 3 days of leptin administration. VMH-injected animals fully recovered from leptin-induced effects within 3 days. We conclude that small doses of leptin that do not effect eating behavior when delivered to the ventricle or the dorsal raphe (another brain region believed to regulate feeding), suppress food intake when injected into the VMH. These data suggest that the VMH or a brain region in close proximity to it is a key target for the biological actions of leptin.

Animals↗
PubMed1995

Intra-VTA injections of the mu-opioid antagonist CTOP enhance locomotor activity.

In this paper we report on the effects of microinjections of the mu-opioid antagonist CTOP (D-Pen-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr-NH2) into the ventral tegmental area (VTA) on activity and ingestive behavior in the rat. Intra-VTA CTOP (0.015, 0.15, and 1.5 nmol per side) dose-dependently increased activity, whereas it had no effect on feeding and drinking behavior. These results are consistent with previous reports that intra-VTA injections of CTOP enhance extracellular dopamine levels in the nucleus accumbens. Furthermore, we propose a model of VTA mu-opioid mechanisms that might account for these surprising effects of intra-VTA CTOP.

Analysis of Variance↗
PubMed1995

Azithromycin for empirical treatment of the nongonococcal urethritis syndrome in men. A randomized double-blind study.

OBJECTIVE: To evaluate the use of single-dose azithromycin for empirical treatment of nongonococcal urethritis. DESIGN: Randomized, double-blind, multicenter trial comparing azithromycin vs doxycycline therapy, with a 2:1 randomization ratio. Patients were evaluated clinically and microbiologically for Chlamydia trachomatis and Ureaplasma urealyticum infection before therapy and at 2 and 5 weeks after study entry. SETTING: Eleven sexually transmitted disease clinics throughout the United States. PATIENTS: A total of 452 men aged 18 years or older with symptomatic nongonococcal urethritis of less than 14 days' duration. INTERVENTION: Patients were treated with either 1.0 g of azithromycin as a single oral dose or 100 mg of doxycycline taken orally twice daily for 7 days. MAIN OUTCOME MEASURES: Clinical resolution of symptoms and signs of nongonococcal urethritis, microbiological cure of C trachomatis and U urealyticum, and occurrence of adverse experiences. RESULTS: Of the 452 patients enrolled, 248 in the azithromycin-treated group and 123 in the doxycycline-treated group were evaluable for clinical response. The two treatment groups were comparable in terms of age, weight, ethnic distribution, sexual preference, sexual activity, and history of prior nongonococcal urethritis or gonorrhea. Sixteen percent of the azithromycin group and 24% of the doxycycline group were culture positive for C trachomatis before therapy, while 38% and 28%, respectively, were culture positive for U urealyticum. The cumulative clinical cure rate was 81% (95% confidence interval [CI], 75% to 85%) in the azithromycin-treated group and 77% (95% CI, 69% to 84%) in the doxycycline-treated group. Clinical cure rates in the two groups were also comparable when patients were stratified by presence or absence of infection with C trachomatis or U urealyticum prior to therapy. Among those infected with C trachomatis, overall microbiological cure rates were 83% (95% CI, 65% to 94%) for azithromycin-treated patients (n = 30) and 90% (95% CI, 68% to 98%) for doxycycline-treated patients (n = 21). Among those infected with U urealyticum, overall microbiological cure rates were 45% (95% CI, 34% to 57%) for azithromycin-treated patients (n = 75) and 47% (95% CI, 30% to 65%) for doxycycline-treated patients (n = 32). Adverse reactions were generally mild to moderate and occurred in 23% of the azithromycin-treated group and 29% of the doxycycline-treated group. CONCLUSIONS: For empirical treatment of the acute nongonococcal urethritis syndrome in men, a single oral dose of azithromycin was as effective as a standard 7-day course of doxycycline in achieving clinical cure. Further, clinical cure rates were comparable with either regimen, regardless of the presence or absence of Chlamydia or Ureaplasma infection.

Adult↗
PubMed1995

Ventral tegmental area opioid mechanisms and modulation of ingestive behavior.

In this paper we report on the effects of intra-VTA infusion of opioid agonists on rat ingestive behavior in a variety of experimental contexts. When the animals were tested outside of their home cages surrounded only by food-pellets (Experiment 1), the injection of the mu-opioid agonist DAMGO, but not the kappa-opioid agonist U-50,488H, into the ventral tegmental area facilitated food-related behaviors, decreasing the latency to feed and increasing the number of interactions with food. When, as in Experiment 2, gnawable objects and a drinking tube were also available, intra-VTA DAMGO gnawing and drinking behaviors, whereas the effects on feeding were negligible. These effects intra-VTA DAMGO increased were greatly enhanced in rats that underwent repeated treatments with amphetamine. On the other hand, when food-related behaviors were studied in a home-cage, where access to the food supply was achieved by entry into a tunnel, latency to feed and total food-intake were not enhanced in tests made during either the dark or the light phase (Experiment 3 and 4). This was true whether powdered standard lab chow or a highly palatable food was available. It appears that when a number of alternative incentive stimuli are available, increases in dopamine transmission such as that induced by intra-VTA DAMGO may ultimately have the effect of interfering with behavior normally directed primarily to one of these stimuli, by enhancing the salience of others. These effects bears some resemblance to the effects of tail-pinch and electrical brain stimulation of the medial forebrain bundle-lateral hypothalamic area on the responses to natural incentive stimuli.

3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cycloh↗
PubMed1995

Elevations of nucleus accumbens dopamine and DOPAC levels during intravenous heroin self-administration.

Extracellular dopamine and DOPAC (3,4-dihydroxyphenylacetic acid) levels in nucleus accumbens were sampled by microdialysis and quantified with high-performance liquid chromatography during intravenous heroin self-administration sessions in rats. Dopamine levels in 10 and 20 min samples were elevated following the first injection of each session, reaching a plateau of elevation within the first two or three injections and falling back toward baseline only when drug access was terminated. Elevations were in the range of 150-300% when unit dosages of 0.05-0.2 mg/kg were given. Increasing the work requirement from FR-1 to FR-10 did not appear to alter the degree of elevation of dopamine levels, and dopamine levels fell during extinction while lever-pressing rates increased 20-fold. While animals compensated for unit dose changes between 0.05 and 0.2 mg/kg/injection, adjusting their response rate such that the same hourly drug intake and the same asymptotic dopamine levels were maintained across these conditions, at 0.4 mg/kg/injection hourly drug intake and asymptotic dopamine levels were elevated beyond the levels sustained by the lower doses. These findings confirm that self-administered doses of intravenous heroin are sufficient to activate the mesolimbic dopamine system and suggest that significant heroin "craving" can emerge when dopamine levels are still moderately elevated, long before the development of dopamine depletion associated with opiate withdrawal.

3,4-Dihydroxyphenylacetic Acid↗
PubMed1995

Chromosomal localization and molecular characterization of 53 cosmid-derived bovine microsatellites.

Gene mapping in cattle has progressed rapidly in recent years largely owing to the introduction of powerful genetic markers, such as the microsatellites, and through advances in physical mapping techniques such as synteny mapping and fluorescence in situ hybridization (FISH). Microsatellite markers are often not physically mapped because they are generally isolated from small insert plasmid libraries, which makes their chromosomal localization inefficient. In this report we describe the FISH mapping of a large group of cosmid-derived bovine microsatellite markers, as our contribution to the European mapping initiative, BovMap. One objective of BovMap is to develop a set of anchored loci for the cattle genome map. Two cosmid libraries were screened with probes corresponding to the (AC)n microsatellite motif. Positive clones were mapped by FISH, and then a subset was further analyzed by sequencing the region flanking the microsatellite repeat. In total, 58 clones were hybridized with chromosomes and identified loci on 22 of the 31 different bovine chromosomes. Three clones contained satellite DNA. Two or more markers were placed on 12 chromosomes. Sequencing of the microsatellites and flanking regions was performed directly from 43 cosmids, as previously reported (Ferretti et al. Anim. Genet. 25, 209-214, 1994). Primers were developed for 39 markers and used to describe the polymorphism associated with the corresponding loci.

Animals↗
PubMed1995

Glucose modulates rat substantia nigra GABA release in vivo via ATP-sensitive potassium channels.

Glucose modulates beta cell insulin secretion via effects on ATP-sensitive potassium (KATP) channels. To test the hypothesis that glucose exerts a similar effect on neuronal function, local glucose availability was varied in awake rats using microdialysis in the substantia nigra, the brain region with the highest density of KATP channels. 10 mM glucose perfusion increased GABA release by 111 +/- 42%, whereas the sulfonylurea, glipizide, increased GABA release by 84 +/- 20%. In contrast, perfusion of the KATP channel activator, lemakalim, or depletion of ATP by perfusion of 2-deoxyglucose with oligomycin inhibited GABA release by 44 +/- 8 and 45 +/- 11%, respectively. Moreover, the inhibition of GABA release by 2-deoxyglucose and oligomycin was blocked by glipizide. During systemic insulin-induced hypoglycemia (1.8 +/- 0.3 mM), nigral dialysate GABA concentrations decreased by 49 +/- 4% whereas levels of dopamine in striatal dialysates increased by 119 +/- 18%. We conclude that both local and systemic glucose availability influences nigral GABA release via an effect on KATP channels and that inhibition of GABA release may in part mediate the hyperexcitability associated with hypoglycemia. These data support the hypothesis that glucose acts as a signaling molecule, and not simply as an energy-yielding fuel, for neurons.

Adenosine Triphosphate↗
PubMed1995

Interstitial fluid concentrations of glycerol, glucose, and amino acids in human quadricep muscle and adipose tissue. Evidence for significant lipolysis in skeletal muscle.

To determine the relationship between circulating metabolic fuels and their local concentrations in peripheral tissues we measured glycerol, glucose, and amino acids by microdialysis in muscle and adipose interstitium of 10 fasted, nonobese human subjects during (a) baseline, (b) euglycemic hyperinsulinemia (3 mU/kg per min for 3 h) and, (c) local norepinephrine reuptake blockade (NOR). At baseline, interstitial glycerol was strikingly higher (P < 0.0001) in muscle (3710 microM) and adipose tissue (2760 microM) compared with plasma (87 microM), whereas interstitial glucose (muscle 3.3, fat 3.6 mM) was lower (P < 0.01) than plasma levels (4.8 mM). Taurine, glutamine, and alanine levels were higher in muscle than in adipose or plasma (P < 0.05). Euglycemic hyperinsulinemia did not affect interstitial glucose, but induced a fall in plasma glycerol and amino acids paralleled by similar changes in the interstitium of both tissues. Local NOR provoked a fivefold increase in glycerol (P < 0.001) and twofold increase in norepinephrine (P < 0.01) in both muscle and adipose tissues. To conclude, interstitial substrate levels in human skeletal muscle and adipose tissue differ substantially from those in the circulation and this disparity is most pronounced for glycerol which is raised in muscle as well as adipose tissue. In muscle, insulin suppressed and NOR increased interstitial glycerol concentrations. Our data suggest unexpectedly high rates of intramuscular lipolysis in humans that may play an important role in fuel metabolism.

Adipose Tissue↗
PubMed1994

Long-term gene expression and phenotypic correction using adeno-associated virus vectors in the mammalian brain.

Adeno-associated viral (AAV) vectors are non-pathogenic, integrating DNA vectors in which all viral genes are removed and helper virus is completely eliminated. To evaluate this system in the post-mitotic cells of the brain, we found that an AAV vector containing the lacZ gene (AAVlac) resulted in expression of beta-galactosidase up to three months post-injection in vivo. A second vector expressing human tyrosine hydroxylase (AAVth) was injected into the denervated striatum of unilateral 6-hydroxydopamine-lesioned rats. Tyrosine hydroxylase (TH) immunoreactivity was detectable in striatal neurons and glia for up to four months and we also found significant behavioural recovery in lesioned rats treated with AAVth versus AAVlac controls. Safe and stable TH gene transfer into the denervated striatum may have potential for the genetic therapy of Parkinson's disease.

Adenoviruses, Human↗