Activity levels after pulmonary rehabilitation - what really happens?

OBJECTIVES: To assess the changes in physical activity in subjects with chronic obstructive pulmonary disease over 6months after pulmonary rehabilitation. DESIGN: Prospective, observational study. Activity was measured over 2-day periods at the end of rehabilitation, and repeated every 6weeks for 6months using the ActivPAL uni-axial accelerometer. These results were compared with the shuttle walking test (SWT) and the St. George's Respiratory Disease Questionnaire (SGRDQ). SETTING: UK community hospital. PARTICIPANTS: Adults completing a community rehabilitation programme. MAIN OUTCOME MEASURE: Time spent standing and mobilising ('uptime'). RESULTS: Of 34 subjects recruited, 28 completed the 6-month study period (mean age 69years, mean forced expiratory volume in 1second 1.3l). Participants wore the monitor for 13.8 to 14.2hours/day. At baseline (post-rehabilitation), participants spent 1.7 [standard deviation (SD) 1.3]hours/day walking and 3.5 (SD 2.6)hours/day standing. Taking the group as a whole, mean uptime decreased marginally by 13.6minutes after 24weeks compared with baseline, with significant individual variability. In all but one subject who showed decreased activity, this was apparent after 6weeks. There were no significant changes in the mean SWT or SGRDQ. Significant associations between total uptime and the SWT were found, but coefficients were weak. It was not possible to predict individual responses from baseline data. CONCLUSION: The accelerometer provides useful supplementary data in patients completing rehabilitation programmes, and the results reveal wide variation. The weak associations between activity data and the SWT suggest that monitors provide additional information. More work is required to determine the factors associated with early deterioration in activity in order to design appropriate interventions.

The incremental shuttle walking test in elderly people with chronic airflow limitation.

BACKGROUND: There is a concern that comorbidity or frailty in older people could limit the usefulness of currently available exercise tests for chronic lung disease. This study evaluated the feasibility and reproducibility of the incremental shuttle walking test (SWT) in people aged 70 years or over, compared exercise tolerance with other disability markers, and assessed whether the SWT is responsive to change after bronchodilators. METHODS: Fifty elderly patients with chronic airflow limitation (CAL) and 32 controls without airflow limitation attempted the SWT before and after combined nebulised salbutamol/ipratropium bromide. Subjects also completed the Nottingham Extended Activities of Daily Living index (NEADL) and the London Handicap score (LHS). RESULTS: Forty four subjects with CAL (88%) and 29 controls (84%) completed the SWT, including many with co-morbidities. Two week repeatability was good and the SWT was strongly associated with EADL (r=0.51, p<0.001) and LHS (r=0.43, p<0.004), but only weakly with forced expiratory volume in 1 second (FEV1) (r=0.31, p=0.05). Subjects with CAL walked a mean distance of 177.7 m compared with 243.3 m in controls (p<0.001); following bronchodilator therapy the distance walked increased in the CAL group by 13.2% (p=0.009). CONCLUSION: The SWT is a feasible and reproducible measure of exercise tolerance in elderly people with and without airflow obstruction and correlates with other markers of disability. It is sensitive to change following bronchodilation in subjects with CAL, although the change correlates less well with improvements in FEV1. Overall, these results suggest that the SWT might be an appropriate measure to assess interventions in elderly people.

Developmental sensitivity of associative learning to cholesterol synthesis inhibitors.

Patients with Smith-Lemli-Opitz syndrome, a genetic disorder associated with severe mental retardation, are unable to convert 7-dehydrocholesterol to cholesterol. Treatment of rats with agents that block cholesterol synthesis produces a sterol profile reminiscent of Smith-Lemli-Opitz patients i.e., low levels of cholesterol accompanied by the appearance of its immediate precursor 7-dehydrocholesterol. In previous work, chronic inhibition of cholesterol synthesis in just-weaned rats impaired acquisition of the classically conditioned eyeblink response. The present study had two primary goals--(1) to determine whether the learning impairment depended on the age in which treatment was initiated; and (2) to determine whether the deficit was associative or due to performance factors. Consistent with earlier work, acquisition of the eyeblink conditioned response was impaired when the 30-day treatment was initiated on postnatal day (PND) 21. Reactivity to acoustic stimuli and to eyelid stimulation were normal, suggesting that the learning impairment was associative in nature. The learning impairment was transitory; acquisition was normal when evaluated 30 days after the cessation of treatment. When treatment was initiated 30 days after weaning (PND 51), acquisition of the eyeblink response was normal. However, brain sterols of young adult rats were less affected than those of just-weaned rats. Thus, there is a developmental sensitivity to cholesterol synthesis blocking agents both in terms of their effects on brain sterols and new motor learning.

Apolipoprotein E is a putative autocrine regulator of the rat ovarian theca cell compartment.

Apolipoprotein (apo) E inhibits androgen production by ovarian theca cells. We found that apo E, as a synthetic peptide mimicked the full-size protein, induced theca and interstitial cell (TIC) apoptosis indicated by pyknotic cell morphology, increased DNA end-labeling (TUNEL), and DNA ladders. None of the low-density lipoprotein (LDL) receptor superfamily members were involved because the universal antagonist of these receptors, receptor-associated protein (RAP), did not block apo E-induced apoptosis. Furthermore, several apo E synthetic peptides that do not bind the LDL receptor did induce TIC apoptosis. Similar to apo E, apoptogenic agents such as ceramide and LY 294002, a phosphatidylinositol (PI) 3-kinase inhibitor, induced apoptosis and suppressed androstenedione production. However, apoptosis alone was not responsible for apo E suppression of androstenedione production because both insulin and IGF-I prevented apo E-induced apoptosis, but neither restored androstenedione production. Theca cells of atretic follicles express the greatest apo E mRNA, and here we show that cultured TIC produce apo E. When considered with the observation of TUNEL-positive theca cells in atretic follicles these results support our hypothesis that intraovarian apo E controls theca cell production of androgen as well as limiting the size of the theca cell compartment.

Is there a relationship between 3-hydroxy-3-methylglutaryl coenzyme a reductase activity and forebrain pathology in the PKU mouse?

Previous reports have suggested that elevated levels of phenylalanine inhibit cholesterol synthesis. The goals of this study were to investigate if perturbations in cholesterol synthesis exist in the PAH(enu2) genetic mouse model for phenylketonuria (PKU), and if so, initiate studies determining if they might underlie the white matter pathology that exists in PKU forebrain. Gross sections and electron microscopy showed that select tracts were hypomyelinated in adult PKU mouse forebrain but not hindbrain. The activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR), the rate controlling enzyme in the cholesterol biosynthetic pathway, was examined in isolated microsomes from forebrain, hindbrain, and liver to assess if perturbations in cholesterol biosynthesis were occurring. HMGR activity was normal in unaffected PKU hindbrain and was increased 2-4-fold in PKU liver compared to control. HMGR activity in the forebrain, however, was decreased by 30%. Because normal numbers of MBP-expressing glia (oligodendrocytes) were present, but the number of glia expressing HMGR was reduced by 40% in the hypomyelinated tracts, the decreased HMGR activity seemed to result from a down-regulation of HMGR expression in affected oligodendrocytes. Exposure of an oligodendrocyte-like glioma cell line to physiologically relevant elevated levels of Phe resulted in a 30% decrease in cholesterol synthesis, a 28% decrease in microsomal HMGR activity, and a 28% decrease in HMGR protein levels. Measurement of HMGR activity after addition of exogenous Phe to control brain microsomes revealed that Phe is a noncompetitive inhibitor of HMGR; physiologically relevant elevated levels of exogenous Phe inhibited HMGR activity by 30%. Taken together, these data suggest that HMGR is moderately inhibited in the PKU mouse. Unlike other cell types in the body, a subset of oligodendrocytes in the forebrain seems to be unable to overcome this inhibition. We speculate that this may be the cause of the observed pathology in PKU brain.

GFAP-positive and myelin marker-positive glia in normal and pathologic environments.

The data herein demonstrate that in addition to the well-characterized myelin marker-positive, glial fibrillary acidic protein (GFAP)-negative, membrane sheet-bearing oligodendrocytes, another type of myelin marker-positive, process-bearing glia exists in normal and pathologic conditions. This second type of myelin marker-positive glia expresses GFAP, and therefore these cells have been referred to as mixed phenotype glia. Although mixed phenotype glia have been documented previously, their identity and function have remained a mystery. The goal of this immunocytochemical study was to further characterize these cells. Using the MBPlacZ transgenic mouse in which beta-galactosidase is under the control of the myelin basic protein (MBP) gene promoter, GFAP-positive/beta-galactosidase-positive and myelin/oligodendrocyte-specific protein (MOSP)-positive/beta-galactosidase-positive cells were detected in subcortical white matter and in perivascular locations within cerebral white and gray matter. In cultures prepared from highly enriched myelin marker-positive immature glia, mixed phenotype glia were detected that were GFAP-positive and either MOSP-, MBP-, O1-, and O4-positive. The expression of multiple myelin markers by mixed phenotype glia may suggest that these cells are of oligodendrocyte origin. Increased numbers of MOSP-positive/GFAP-positive mixed phenotype glia were detected in sections from adult hypomyelinated brain from shiverer, quaking, and PKU mice compared to myelinated control adult mouse brain. Similarly, cultures from control brain exposed to elevated pH for 2-3 weeks showed dramatically increased numbers of mixed phenotype glia (80%) compared to control (<10%). Increased numbers of mixed phenotype glia also were detected in shiverer cultures (40%). Since increases in the number of mixed phenotype glia occur in shiverer, quaking, and PKU mouse brain, these data suggest that mixed phenotype glia contribute to gliosis in pathologic white matter.

Redistribution of cholesterol in oligodendrocyte membrane sheets after activation of distinct signal transduction pathways.

Cultured oligodendrocytes produce extensive membrane sheets that contain an internal lacy network of vein-like structures composed of microtubules, actin filaments, and 2'3'-cyclic nucleotide 3'-phosphohydrolase (CNPase). These cytoplasmic vein-like structures surround domains of myelin basic protein (MBP). Using the antibiotic filipin, that binds to cholesterol, the relationship between plasma membrane cholesterol and cytoskeleton in membrane sheets was examined. Our results show that cholesterol was relatively uniformly distributed within the plasma membranes of prefixed control oligodendrocyte membrane sheets. When live cultures were extracted with Triton X-100, however, a subpopulation of cholesterol molecules remained colocalized with cytoskeleton in the membrane sheets. Activation of two well-characterized signaling pathways that differentially affect microtubule and actin filament stability in membrane sheets resulted in an apparent massive lateral movement of cholesterol molecules away from membrane regions overlying internal MBP domains to membrane tracts directly overlying cytoplasmic cytoskeletal veins. Depolymerization of microtubules by colchicine resulted in redistribution of cholesterol directly over actin filaments, whereas depolymerization of actin filaments by cytochalasin B resulted in redistribution of cholesterol directly over CNPase/microtubular veins. These data suggest that cholesterol forms an association with cytoskeletal components or proteins associated with cytoskeleton. These data also suggest that cholesterol, via interactions with cytoskeleton, plays a role in signaling pathways in oligodendrocyte membrane sheets.

Quality of life in elderly subjects with a diagnostic label of asthma from general practice registers.

The aim of this study was to assess health-related quality of life (QoL) in elderly subjects with a diagnostic label of asthma from a general practice population, and to determine the main contributory factors. Sixty people aged > or =70 yrs with a primary care diagnostic label of asthma, and 43 control subjects were recruited. Assessment of bronchodilator response, and oral steroid trials were conducted where possible. The main outcome measures were QoL scores for the Short Form (SF)-36 and the St George's Respiratory Questionnaire (SGRQ). In the asthma group, 29 subjects demonstrated a significant airway response to bronchodilators or steroids. Mean SF-36 scores were significantly worse in the total asthma group for components of physical function, physical role limitation, and general health, although psychological scores were similar. QoL remained worse than controls in those subjects with a significant bronchodilator response. Dyspnoea and depression accounted for 61% of the variance in the SGRQ, but forced expiratory volume in one second was not an independent variable. Quality of life is impaired in elderly people with a diagnosis of asthma, including those with demonstrable airway variability. Many older subjects with asthma note a variety of symptoms, highlighting the need for further research into the adequacy and efficacy of their treatment.

Apolipoprotein E peptide stimulation of rat ovarian theca cell androgen synthesis is mediated by members of the low density lipoprotein receptor superfamily.

Ovarian androgen production is rate limiting for follicular maturation and can induce follicular atresia. Thus, it is important to define the actions of the intraovarian agents, such as apolipoprotein (apo) E, that modulate theca cell androgen production. Theca cell androgen production is stimulated at low concentrations and inhibited at higher concentrations of native apo E. The apo E peptide, acetyl-Y(LRKLRKRLLRDADDL)(2)C or acetyl-Y(141-155)(2)C, has low density lipoprotein (LDL) receptor and LDL receptor-related protein-binding activity, and it mimics the activity of native apo E in the theca-interstitial cell system. To define the role of members of the LDL receptor superfamily in the apo E peptide-mediated responses, we found that receptor-associated protein prevented the stimulation without altering the inhibition of androstenedione production. The apo E peptide (129-162), which has no LDL receptor-binding activity, did not stimulate androstenedione production. The apo E peptide acetyl-Y(141-155)(2)C did not stimulate androstenedione production when cell surface heparan sulfate proteoglycans were degraded with heparinase. The apo E peptide acetyl-Y(141-155)(2)C bound to heparin, a property of LDL receptor ligands, and in this complex the peptide had no effect on androstenedione production. These observations support the conclusion that apo E-mediated stimulation, but not inhibition, of ovarian theca cell androstenedione production was mediated by members of the LDL receptor superfamily.

A syngeneic mouse glioma model for study of glioblastoma therapy.

Animal models of human tumors serve a vital role in the development and testing of new anticancer therapies. Since the immune system is likely to play an essential role in tumor eradication, there is a particular need for modeling human disease in immunocompetent hosts. Few models of glioma have been developed in immunocompetent mice that are commercially available and none of these tumors have histological and antigenic characteristics of human gliomas. We have used a cell line, 4C8, derived from a spontaneous glioma-like tumor that arose in a transgenic mouse to develop a new glioma model. The intracranial injection of 4C8 cells into immunocompetent syngeneic B6D2F1 mice resulted in tumors that were densely cellular, developed a pseudopallisading pattern of necrosis, and expressed GFAP; all important features of human malignant gliomas. The average neurological endpoint was 51 days after intracranial injection. The 4C8 cells also grew rapidly in the flank, retaining histologic features seen in intracranial tumors. Flank tumors reached an average volume of 100 mm3, a volume ideal for therapy testing, by 34 days postinjection. These results suggest that the 4C8 mouse glioma model is an excellent system in which to test new antiglioma therapies for use in humans.

An apolipoprotein E synthetic peptide selectively modulates the transcription of the gene for rat ovarian theca and interstitial cell P450 17alpha-hydroxylase, C17-20 lyase.

Ovarian theca/interstitial cells produce androgens in response to luteinizing hormone (LH) stimulation and apolipoprotein (apo) E exerts a selective effect on the type of steroid product made by these cells. We have identified an apoE synthetic peptide containing the low density lipoprotein (LDL) receptor binding domain, acetyl-Y(LRK LRKRLLRDADDL)2C, that mimics the activity of native apoE. Depending on the concentration, the apoE synthetic peptide either enhanced or inhibited the LH-stimulated production of androstenedione with concomitant changes in the mRNA for its synthetic enzyme, P450 17alpha-hydroxylase, C17-20 lyase, without any changes in progesterone production or the mRNA for its synthetic enzyme, P450 cholesterol side-chain cleavage. The apoE synthetic peptide caused changes in the rate of transcription of the mRNA for P450 17alpha-hydroxylase, C17-20 lyase without altering its stability. Pretreatment of the theca/interstitial cells with receptor-associated protein, which blocks apoE binding to members of the LDL receptor superfamily, prevented the apoE synthetic peptide-mediated stimulation of androstenedione and mRNA for P450 17alpha-hydroxylase, C17-20 lyase, but did not attenuate the inhibitory activity of the peptide. Thus, apolipoprotein E selectively altered the type of steroid made by ovarian theca/interstitial cells by regulating the transcription of mRNA for the gene for P450 17alpha-hydroxylase, C17-20 lyase, in part through its interaction with apolipoprotein E-specific receptors of the LDL receptor superfamily.

Risk-factor assessment for falls: from a written checklist to the penless clinic.

OBJECTIVE: to audit risk-factor identification of fallers before and after an education programme and the insertion of a written checklist in medical notes. Risk-factor identification in a dedicated, computerized falls clinic was then examined. METHODS: documentation of risk factors for falls was studied on wards and a self-auditing 'penless' clinic for fallers subsequently set up to generate reports for medical notes and letters for general practitioners. RESULTS: risk-factor identification improved after the insertion of the checklist but remained relatively poor. A dedicated clinic allowed almost complete identification of risk factors. Of the first 112 patients (median age 82) seen in the clinic, 75 (67%) were housebound. Remediable risk factors--e.g. inappropriate medication (67%), unsatisfactory footwear (59%) and postural hypotension (17%)--were found in most. Thirty-three patients (29%) had difficulty with alarm raising. CONCLUSION: ward-based intervention showed limited capacity to identify risk factors for falls: a dedicated clinic was more successful. The use of a portable computer with a programme to screen fallers for risk factors is worthy of consideration.

Cardiovascular defects among the progeny of mouse phenylketonuria females.

In a genetic mouse model of human phenylketonuria we have examined the offspring of hyperphenylalaninemic mothers for the presence of cardiovascular defects, an important feature of the pathology of the human maternal phenylketonuria syndrome. Beginning at 14.5 d after conception (75% through gestation), a variety of cardiovascular defects became apparent among the progeny of the hyperphenylalaninemic females. These defects ranged from mild to serious and correlated with the maternal but not the fetal Pah genotype. Nearly all of the defects were vascular, however, whereas the most reported in humans so far have been cardiac. The predisposing biochemical condition in this mouse disease model seems to be the same as in the human disease; elevated maternal blood phenylalanine levels concentrated across the placental barrier to produce a teratogenic developmental environment. This model for congenital cardiovascular defects should enhance two related areas of research. 1) It should allow a more thorough investigation of the relationship between maternal diet and maternal phenylketonuria birth defects, and 2) it should provide an experimental tool to gain insight into the normal process of cardiovascular development.

A hospital-based case-control study of quality of life in older asthmatics.

Asthma is a common but neglected problem in older people, the impact of which is relatively unstudied. The aim of this study was to objectively assess quality of life and depression in older asthmatics. The subjects studied were 50 hospitalized known asthmatics, over 55 yrs of age (mean age 72 yrs). Of these, 40 had objective evidence of asthma, and were compared to 40 age- and sex-matched controls. Using a structured questionnaire, the Geriatric Depression Score, subjective health status (short form (SF)-36), and other comparative disability data were recorded. Spirometric results were also recorded. Depressive symptoms were common in both groups but were not significantly different. Mean SF-36 scores were significantly worse in the asthmatics, especially for components of physical function (p = 0.04), physical role limitation (p = 0.01), energy (p = 0.01), health change (p = 0.01), and general health perception (p = 0.01). However mental, mental role and social scores were similar in both groups. We conclude that quality of life is impaired in hospitalized asthmatics compared to controls. Physical components appear to be most adversely affected. Depressive symptoms are common but no overall difference was found for psychological disability. Older asthmatics appear to adapt well to adverse situations.

Regulation of cytoskeleton by myelin components: studies on shiverer oligodendrocytes carrying an Mbp transgene.

Oligodendrocytes from the shiverer mutant mouse are missing most of the myelin basic protein (Mbp) gene. In axon-free cultures, they produce membrane sheets with abnormally assembled microtubule and actin-based structures. This suggests that an Mbp gene product may have an important role in regulating the organization and stability of the wild-type oligodendrocyte cytoskeleton. We now present evidence extending these observations, using cultured oligodendrocytes that carry both the shiverer mutation and the Mbp1 transgene which partially corrects their deficit. Shiverer oligodendrocytes that carry one dose of the Mbp1 transgene abnormally express MBP along major cytoskeletal vein-like structures in processes and sheets. Shiverer oligodendrocytes that carry two doses of the Mbp1 transgene contain two types of membrane sheet regions, i.e. regions filled with aberrant punctate foci of MBP, and regions with normal domains of MBP. Immunocytochemical staining data show that the distribution of cytoskeleton and associated 2',3'-cyclic nucleotide 3'-phosphohydrolase (CNPase) is dependent upon how MBP is organized. Bundling of actin filaments occurs only around MBP domains, and the colocalization of CNPase along microtubular structures also appears to be regulated by MBP domains in sheets. Multinucleated oligodendrocytes are observed, a likely result of the inability of dividing pro-oligodendrocytes to bundle actin filaments. In addition, the ability of MBP to mediate extracellular signals that modulate cytoskeleton appears to be dependent upon MBP's organization. Transduction of the galactocerebroside signaling pathway, which results in the destabilization of microtubules but not actin filaments, occurs only in sheets containing MBP domains. The distribution of MBP, however, does not affect the myelin/oligodendrocyte-specific protein signaling pathway, which results in growth of microtubular structures and extensive destabilization of the actin cytoskeleton.

Evidence for central nervous system glial cell plasticity in phenylketonuria.

Phenylketonuria (PKU) is caused by mutation(s) in the phenylalanine hydroxylase (PAH) gene which lead to deficient PAH activity and an accumulation of phenylalanine in the blood. The primary pathologic finding is hypomyelination and gliosis of central nervous system white matter. Similar white matter pathology is observed in the Pahenu2 mouse, a genetic model for PKU. We studied this mouse to examine the basis for these neuropathologic changes in PKU and to determine if hypomyelination and gliosis occur independently or are interrelated. Although white matter tracts within PKU brains are hypomyelinated, immunostaining and Western blot analyses revealed that these tracts contain abundant amounts of myelin markers, i.e. myelin basic protein (MBP), 2',3'-cyclic nucleotide 3'phosphohydrolase, and myelin/oligodendrocyte-specific protein (MOSP). However, Western blot analyses also showed that MBP isoform expression was aberrant. Investigation of individual cells was performed by extraction of tissue sections with Triton X-100. Most of the MOSP was extracted, with the remaining MOSP clearly visible in dual labeled cells, i.e. MOSP was colocalized along glial fibrillary acidic protein (GFAP) filaments. Cells expressing both MBP and GFAP were also identified in optic tract. Double labeling with a riboprobe for MBP and antibodies specific for GFAP revealed that the majority of GFAP-positive cells expressed MBP mRNA. Our in vitro studies examined the response of cultured wild type oligodendrocytes to elevated phenylalanine for 4 weeks (wk). Under these conditions, about 50 these conditions, about 50% of the oligodendrocytes expressed GFAP filaments and failed to elaborate membrane sheets. Proliferation of astrocytes appears not to be the source of gliosis, since the nuclei of GFAP-positive cells in the PKU brains did not immunostain for proliferating cell nuclear antigen. Dual-labeled cells were detected in normal mouse brain sections; however, PKU mouse white matter tracts were found to contain about twice the number of dual-labeled cells compared to normal tissue. Taken together, these data suggest that both myelinating and nonmyelinating oligodendrocytes are present in the normal adult brain, and that in response to a toxic factor such as elevated phenylalanine, myelinating oligodendrocytes adopt a nonmyelinating phenotype that expresses GFAP. Since myelinating Schwann cells and GFAP-positive nonmyelinating Schwann cells are normally present in adult peripheral nerve, and the myelinating Schwann cells react to pathologic situations by switching to GFAP-positive nonmyelinating cells, it may be that oligodendrocytes and Schwann cells are more similar than previously thought.

Comparison of promoter strengths on gene delivery into mammalian brain cells using AAV vectors.

Recent reports have suggested that delivery of genes flanked by AAV ITRs may be useful for gene therapy of diseases that involve the brain. We have compared the efficiency of gene expression in vitro in CNS-derived cells from four different promoters when the transgene is flanked by AAV ITRs, using both transfection via cationic liposomes, and infection via rAAV. The human cytomegalovirus (CMV) immediate-early enhancer/promoter, the SV40 early enhancer/promoter, the JC polymovirus promoter, and the chicken beta-actin promoter coupled to the CMV enhancer were able to drive expression of the reporter gene beta-galactosidase in all tumor and primary brain cell cultures tested. Although the relative order of efficiency differed between cell types, the CMV promoter was always the strongest, generally by at least one order of magnitude. A comparison of the relative levels of expression seen between different cell types on transfection and infection suggest that not all CNS-derived cells are infected equally efficiently by rAAVs. High level of expression were seen within 24 h of transgene delivery by either transfection or infection, dropping dramatically within days. All cell types and promoters showed the same decline, suggesting that transient expression by rep-rAAVs may be efficient, but stable expression as detected in this system is a low frequency event. In vivo studies using the CMV promoter also suggest that although rep-rAAVs are able to infect efficiently CNS cells and produce high levels of gene expression shortly after transduction, the majority of such infections do not lead to stable high-level expression of transgenes.