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Biomedical subjects

N Provost

Publications and source records attributed to N Provost.

At least 19 recordsLinked to original sources
PubMed2006

Restoration of vision in RPE65-deficient Briard dogs using an AAV serotype 4 vector that specifically targets the retinal pigmented epithelium.

Previous studies have tested gene replacement therapy in RPE65-deficient dogs using recombinant adeno-associated virus 2/2 (rAAV2/2), -2/1 or -2/5 mediated delivery of the RPE65 gene. They all documented restoration of dark- and light-adapted electroretinography responses and improved psychophysical outcomes. Use of a specific RPE65 promoter and a rAAV vector that targets transgene expression specifically to the RPE may, however, provide a safer setting for the long-term therapeutic expression of RPE65. Subretinal injection of rAAV2 pseudotyped with serotype 4 (rAAV2/4) specifically targets the RPE. The purpose of our study was to evaluate a rAAV2/4 vector carrying a human RPE65cDNA driven by a human RPE65 promoter, for the ability to restore vision in RPE65-/- purebred Briard dogs and to assess the safety of gene transfer with respect to retinal morphology and function. rAAV2/4 and rAAV2/2 vectors containing similar human RPE65 promoter and cDNA cassettes were generated and administered subretinally in eight affected dogs, ages 8-30 months (n = 6 with rAAV2/4, n = 2 with rAAV2/2). Although fluorescein angiography and optical coherence tomography examinations displayed retinal abnormalities in treated retinas, electrophysiological analysis demonstrated that restoration of rod and cone photoreceptor function started as soon as 15 days post-injection, reaching maximal function at 3 months post-injection, and remaining stable thereafter in all animals treated at 8-11 months of age. As assessed by the ability of these animals to avoid obstacles in both dim and normal light, functional vision was restored in the treated eye, whereas the untreated contralateral eye served as an internal control. The dog treated at a later age (30 months) did not recover retinal function or vision, suggesting that there might be a therapeutic window for the successful treatment of RPE65-/- dogs by gene replacement therapy.

Animals↗
PubMed2001

Immediate and long-term safety of recombinant adeno-associated virus injection into the nonhuman primate muscle.

Previous studies on distribution and toxicity of viral vectors administered in monkeys indicated that the nonhuman primate model has a reasonable predictive value for clinical applications. In this study, eight macaques were injected intramuscularly with recombinant adeno-associated virus (rAAV) at doses similar to those administered to hemophilia B patients, and followed to analyze the dissemination and shedding in biological samples and long-term persistence in distant organs. Following rAAV delivery, we found vector genome in various biological fluids for up to 6 days and infectious particles exclusively in the serum during the first 48-72 hours. rAAV sequences were detected in peripheral blood mononuclear cells (PBMC) for up to 10 months. At necropsy, 8 to 18 months after rAAV delivery, rAAV sequences were found in lymph nodes and livers but never in the gonads. Tissue examination, of liver in particular, showed no abnormalities. We concluded that during our experimental time frame, rAAV-mediated gene transfer into skeletal muscle of macaques seemed to be safe with respect to the recipient and the environment. However, it was associated with a transient viremia and the persistence of rAAV sequences in PBMC, lymph nodes, and liver, the long-term consequences of which remain unknown.

Animals↗
PubMed2000

Hyaluronidase enhances recombinant adeno-associated virus (rAAV)-mediated gene transfer in the rat skeletal muscle.

Skeletal muscle is a privileged target for long-term rAAV-mediated gene transfer in mouse, rat, dog and non-human primates. Intramuscular injections of rAAV encoding human factor IX in hemophilia B patients have been initiated, based on promising results gathered in affected dogs. We found that intramuscular rAAV administration in rats resulted in restricted transduction essentially along the myofibers axis with poor lateral diffusion. This suggested that the transduction rate might be limited by the ability of the virus to reach sites distant from the injection point. We tested whether hyaluronidase, an enzyme which dissociates the extracellular matrix, could enhance vector diffusion when injected in the rat muscle before administration of rAAV encoding either nuclear-localized beta-galactosidase (rAAVCMVnlsLacZ) or the human alpha-1-antitrypsin (rAAVCMVhAAT) under the control of the cytomegalovirus immediate--early promoter (CMV). The results showed that pretreatment of the rat anterior tibialis muscle with hyaluronidase resulted in: (1) a larger diffusion of the virus indicated by an increase in the area containing LacZ-transduced fibers, and (2) a two- to three-fold increase of transduction efficiency measured by the number of LacZ-positive fibers or by the hAAT serum concentration. We also provide evidence that hyaluronidase was well tolerated and was not associated with short- or long-term toxicity evaluated by morphological studies. Finally, in our experimental conditions, hyaluronidase did not promote rAAV dissemination to other organs as assessed by PCR to detect vector sequences. We conclude that pretreatment of skeletal muscle by hyaluronidase, a clinically available reagent, was harmless and resulted in a consistent and significant increase in rAAV diffusion and transduction levels.

Animals↗
PubMed2000

Comparison of UVA protection afforded by high sun protection factor sunscreens.

UVA protection afforded by 6 different sunscreens with a sun protection factor of 21 or more was compared by means of the persistent pigmentation darkening method. Colorimetric and visual assessment showed significant differences in UV radiation-induced pigmentation at 2 hours. The labeled sun protection factor of the tested sunscreens was not predictive of UVA protection level.

Administration, Topical↗
PubMed1998

Laboratory tests and imaging studies in patients with cutaneous malignant melanoma.

Laboratory tests and imaging studies are often ordered for asymptomatic patients with malignant melanomas (MMs) in an effort to detect subclinical metastatic disease. However, their sensitivity and specificity for detecting cryptic metastases are not well established. A review of the literature on laboratory tests and imaging studies for MM metastases was undertaken to address the usefulness of such investigations in asymptomatic patients with MM in AJCC (American Joint Committee on Cancer system of classification) stages I, II, and III. A review of the pertinent literature since 1966 was conducted through MEDLINE, Medica, and Cancerlit. Laboratory tests and imaging studies revealed occult MM metastases in only a small number of the thousands of reported patients with putative AJCC stage I, II, and III MM. However, for those diagnosed with limited metastases, surgical removal with or without immunotherapy, chemotherapy, or radiotherapy can lead to long-term remissions in some patients. For patients with asymptomatic AJCC stage I or II disease, chest roentgenograms (CXR) and blood lactic dehydrogenase (LDH) levels may be obtained at low cost and prove to be of benefit if metastases are identified. For patients with AJCC stage III disease, computed tomographic (CT) scans of the thorax, abdomen, and pelvis (especially when the primary cutaneous site of the melanoma is below the waist) may be considered for detecting metastatic MM. Other tests, such as magnetic resonance imaging (MRI) scans of the brain, may be ordered based on symptoms or physical findings. In the future, technologically improved techniques and newer methods may prove cost-effective for detecting treatable asymptomatic MM metastases. Furthermore, improvement in treatments will also influence the indications for the search for occult MM metastases. At this time there is a need for an international consensus conference on laboratory tests and imaging studies for occult melanoma metastases.

Clinical Chemistry Tests↗
PubMed1998

Actions of HSVtk and connexin43 gene delivery on gap junctional communication and drug sensitization in hepatocellular carcinoma.

We have previously demonstrated that transfected hepatocellular carcinoma cells (Hepa1-6) with one copy (pAGO) and two copies (pYED) of the HSVtk gene, using liposomes, induced cell death of untransfected cells in the presence of ganciclovir (GCV). This phenomenon is called the 'bystander effect'. To determine whether an elevated level of connexin43 increases the bystander effect, we have cotransfected Hepa1-6 cells with a plasmid containing the HSVtk gene driven by the alpha-fetoprotein promoter (pFTK) or pAGO or pYED and connexin43. The results showed that, after GCV treatment, the percentage of growth inhibition was higher (25-30%) in cells cotransfected with HSVtk and connexin43 than in cells transfected only with HSVtk gene. The IC50 of GCV on cells transfected with pFTK/Connexin43 was 17.85-fold lower than cells transfected with pFTK alone. To improve these results, stable connexin43 transduced Hepa1-6 cells were transfected with pFTK followed by GCV treatment. In this case, the cell growth was markedly inhibited as compared with parental cells. Furthermore, we have studied the correlation between the expression of the HSVtk and the connexin43 proteins. Using flow cytometric analysis, scrape loading/dye transfer and immunoblotting assay we found that the cells transfected separately by pAGO, pYED, pFTK and pLTR-Cx43 showed an increase of connexin43 protein. This study indicates that transfecting Hepa1-6 cells with both connexin43 and HSVtk genes up-regulates connexin43 expression which enhances the bystander effect and subsequently tumor cell death.

Animals↗
PubMed1998

Comparison of conventional photographs and telephonically transmitted compressed digitized images of melanomas and dysplastic nevi.

BACKGROUND: One of the most difficult problems in the in vivo diagnosis of cutaneous tumors is the differentiation clinically between early malignant melanoma (MM) and atypical (dysplastic) melanocytic nevi (AMNs) because these lesions share clinical features. High-quality digital imaging systems and store-and-forward technology have the potential for use in a teledermatology system with which experts would be able to immediately transmit their diagnostic opinions concerning these challenging lesions. OBJECTIVE: The main purpose of this study was to determine if the clinical and dermoscopic diagnoses and the dermoscopic features of AMN and early MM are unaltered after telephonic transmission of their digitized images. METHODS: Conventional and dermoscopic photographic transparencies of 22 AMNs and 9 early MMs, viewed on rearview projectors and then scanned, compressed, transmitted (Internet) and viewed on color monitors, were evaluated. RESULTS: The concordance in the diagnosis of AMN and of early MM by all four observers, both clinically and dermoscopically, when comparing rearview-projected conventional transparency slides to transmitted, compressed, digitized images, was high. For most specific dermoscopic features, the concordance was good, although less so for the presence or absence of some dermoscopic creatures, namely 'dots', 'blue/gray' color and 'red' color. CONCLUSION: The results reported support the conclusion that Internet transmission of digitized images of MMs and AMNs retains sufficient information for diagnostic purposes. This study is a step in the creation of an international teledermoscopy network for pigmented cutaneous lesions.

Computer Communication Networks↗
PubMed1997

A study of large congenital melanocytic nevi and associated malignant melanomas: review of cases in the New York University Registry and the world literature.

BACKGROUND: Patients with large congenital melanocytic nevi (LCMN) are at greater risk for the development of malignant melanoma (MM) than are persons in the general population. OBJECTIVE: Our purpose was to identify the clinical features of LCMN in those patients in whom MMs actually developed. METHODS: The records of 117 patients in the New York University Registry of LCMN and the reports of 172 cases of LCMN in the world literature were studied. RESULTS: Of the 289 cases of LCMN studied, 34 patients (12%) had primary cutaneous MMs within their nevi; in two additional patients, MMs developed at cutaneous sites other than within their nevi. All patients in whom MM developed within LCMN had nevi in axial locations; however, 91% of the LCMN were axial. No MM was found that had arisen in any of the 26 LCMN confined to the extremities. In addition, no MM was found that had arisen in thousands of satellite nevi. CONCLUSION: When MM develops within an LCMN, it generally does so in those LCMN in an axial location. The absence of cases of MM arising in LCMN confined to the extremities suggests that such nevi represent lower risk lesions, but the number of extremity nevi analyzed was too small to allow definitive conclusions. A striking finding was the absence of MMs arising in satellite nevi.

Humans↗
PubMed1997

Laboratory tests and imaging studies in patients with cutaneous malignant melanomas: a survey of experienced physicians.

BACKGROUND: The presence or absence of metastases is important in determining prognosis and treatment options for patients with malignant melanoma (MM). Laboratory tests and imaging studies are ordered for patients with MMs but without symptoms in an effort to detect occult metastases. However, which laboratory tests and imaging studies to order and how often to reorder them is not well established. OBJECTIVE: Our purpose was to determine which tests and studies are ordered by physicians with major responsibilities for the care of patients with MM. METHODS: Physicians were surveyed by questionnaire about the laboratory tests and imaging studies they ordered for MM stages 0, I, II, and III. RESULTS: Of the 35 physicians queried, 30 (86%) responded to the survey. The majority of physicians order tests as follows: no tests for MM in situ; roentgenography of the chest with or without initial lactic acid dehydrogenase/liver function tests for stages I, II, and III and during follow-up for stages IB, II, and III (more frequently as the Breslow thickness increases); and baseline computed tomographic or magnetic resonance imaging scans of the chest, abdomen/pelvis, and brain for stage III. CONCLUSION: Although the pattern of ordering examinations was similar for the majority of respondents, there was significant variability among experienced physicians in ordering laboratory tests and imaging studies in the search for occult metastases in patients with asymptomatic MM. The laboratory tests and imaging studies ordered and their frequency depend on the stage of the MM and sometimes on other risk factors.

Clinical Laboratory Techniques↗
PubMed1997

Presence of integrated DNA sequences of adeno-associated virus type 2 in four cell lines of human embryonic origin.

The human helper virus-dependent parvovirus adeno-associated virus (AAV) has been found in human female genital tissues including material from first trimester miscarriage. In the latter case, AAV type 2 (AAV-2) DNA and viral proteins were detected mainly in the trophoblast cell layer of placenta. In this report, we present evidence that AAV DNA is also present in established human trophoblast cell lines (JEG-3, JAr, BeWo) and in the human amnion cell line FL. In cells of these lines, AAV-2 DNA could be detected both by PCR and Southern blot analysis. Restriction enzyme analysis indicated that AAV DNA was integrated into the host cell genome. Although the cell lines supported AAV replication when infected with AAV-2 and adenovirus type 2 (Ad2) as a helper virus, superinfection with Ad2 alone did not induce replication of AAV DNA, i.e. it failed to rescue AAV from its integrated state. This is probably due to rearrangements within the integrated AAV genome. The presence of AAV DNA in cells derived from human embryonic tissue corroborates the suggestion that human embryonic tissue may be one of the targets of AAV infection.

Cell Line↗
PubMed1996

Neurocutaneous melanosis: clinical features of large congenital melanocytic nevi in patients with manifest central nervous system melanosis.

BACKGROUND: Patients with a large congenital melanocytic nevus (LCMN) may have associated leptomeningeal melanocytosis with or without central nervous system (CNS) melanomas. These patients are considered to have neurocutaneous melanosis, a disorder that, when symptomatic or otherwise manifest neurologically, carries a poor prognosis even in the absence of malignancy. OBJECTIVE: Our purpose was to identify typical clinical features in patients who have manifest CNS melanosis in association with LCMN. METHODS: The records of 117 patients with LCMN in the New York University Registry of LCMN and the reports of 172 cases of LCMN in the world literature were included for features that might signal a high risk for the development of manifest CNS involvement. RESULTS: Of the 289 patients with LCMN, 33 had manifest CNS melanosis. In all 33 in whom symptomatic neurocutaneous melanosis was diagnosed, the LCMNs were present in a posterior axial location on the head, neck, back, and/or buttocks. "Satellite" nevi were known to be present in 31 of the 33 patients. CONCLUSION: Patients with LCMN in a posterior axial location, especially when associated with "satellite" melanocytic nevi, are at greater risk for the development of manifest neurocutaneous melanosis than patients with LCMN limited to the extremities or those who are lacking satellite nevi.

Adolescent↗
PubMed1994

[Study of femoro-tibial cartilage diseases with MRI].

Magnetic resonance imaging provides excellent tissue contrast resolution and can therefore be used to visualize joint cartilage. The goal of this study was to evaluate the value of magnetic resonance imaging for the diagnosis of femorotibial cartilage lesions. Spin-echo magnetic resonance imaging was performed in 37 patients. As compared with arthroscopy, the accuracy, sensitivity, and specificity of magnetic resonance imaging were 72.5%, 47%, and 95.5%, respectively. Only advanced cartilage lesions with exposure of subchondral bone were clearly seen on magnetic resonance sections. Limiting factors included insufficient spatial resolution and the chemical shift phenomenon. Spin-echo sequences used in everyday practice are not appropriate for detecting small cartilage lesions. Controlled studies are needed to determine whether gradient echo sequences are more satisfactory.

Adult↗
PubMed1994

[Fibrofolliculoma. Treatment with copper vapor laser].

There are many clinical presentations to fibrofolliculoma, described by Birt, Hogg and Dubé: the solitary and multiple forms, with or without other skin tumors, could also be markers of intestinal polyposis. Little is known of its pathogenesis. A case of multiple fibrofolliculoma of the face and neck is described. A new therapeutic approach by copper vapour laser is proposed.

Adult↗