[Lower bodylift after massive weight loss: Retrospective study of satisfaction, complications and quality of life. About 76 patients over 4years]. / Bodylift inférieur après amaigrissement massif : étude rétrospective de la satisfaction, des complications et de la qualité de vie. À propos de 76 patients sur 4ans.

INTRODUCTION: Massive weight loss leads to important cutaneous deformities with physical and psychological repercussions for patients. Lower bodylift is a procedure, which can restore the body contour. The aim of this study is to evaluate satisfaction and quality of life in patients who underwent lower bodylift and to review the complications of these procedures. MEANS AND METHODS: This is a retrospective study of 76 patients who underwent lower bodylift between 2012 and 2016. We reviewed the complications of these procedures. Satisfaction and quality of life were assessed using Body-QoL questionnaire. RESULTS: Seventy-six patients were included with a mean age of 39.2years. The average body mass index was 27.6kg/m2 with a mean weight of 71.2kg at the time of surgery and a mean weight loss of 48.6kg. Twenty-three patients developed one complication: 22 minor and 1 major. Forty-eight patients answered the questionnaire. Satisfaction was rated "very good" by 41 patients (85.4%) and "good" by 5 patients (10.4%). The Body-QoL questionnaire's analysis showed an improvement of quality of life socially, sexually, in the body regard and in physical symptoms. CONCLUSIONS: Lower bodylift is the only procedure, which can restore circumferential body contour. Despite the minor complications reviewed, the degree of satisfaction of the patients is very high. The quality of life of these patients, after massive weight loss, is also highly improved by these procedures. With the worldwide development of obesity and bariatric surgery, this study demonstrated that the operation should be proposed to patients with massive weight loss to improve quality of life.

Sustained inhibition of hepatitis B virus replication in vivo using RNAi-activating lentiviruses.

Chronic infection with hepatitis B virus (HBV) puts individuals at high risk for complicating cirrhosis and liver cancer, but available treatment to counter the virus rarely eliminates infection. Although harnessing RNA interference (RNAi) to silence HBV genes has shown the potential, achieving efficient and durable silencing of viral genes remains an important goal. Here we report on the propagation of lentiviral vectors (LVs) that successfully deliver HBV-targeting RNAi activators to liver cells. Mono- and tricistronic artificial primary microRNAs (pri-miRs) derived from pri-miR-31, placed under transcriptional control of the liver-specific modified murine transthyretin (mTTR) promoter, caused efficient inhibition of HBV replication markers. The tricistronic cassette was capable of silencing a mutant viral target and the effects were observed without disrupting the function of an endogenous miR (miR-16). The mTTR promoter stably expressed a reporter transgene in mouse livers over a study period of 1 year. Good silencing of HBV genes, without evidence of toxicity, was demonstrated following intravenous injection of LVs into neonatal HBV transgenic mice. Collectively, these data indicate that LVs may achieve sustained inhibition of HBV replication that is appealing for their therapeutic use.

In the rat liver, Adenoviral gene transfer efficiency is comparable to AAV.

Adenoviral (AdV) and Adenovirus-associated viral (AAV) vectors both are used for in vivo gene therapy of inherited liver disorders, such as Crigler-Najjar syndrome type 1. In a relevant animal model, the Gunn rat, both vectors efficiently correct the severe hyperbilirubinemia characteristic of this liver disorder. Although the clinical use of AAV is more advanced, as demonstrated by the successful phase 1 trial in hemophilia B patients, because of its large cloning capacity AdV remains an attractive option. A direct comparison of the efficacy of these two vectors in the liver in a relevant disease model has not been reported. Aim of this study was to compare the efficiency of clinically applicable doses of both vectors in the Gunn rat. AdV or scAAV (self-complimentary AAV) ferrying identical liver-specific expression cassettes of the therapeutic gene, UGT1A1, were injected into the tail vein. As the titration methods of these two vectors are very different, a comparison based on vector titers is not valid. Therefore, their efficacy was compared by determining the amount of vector genomes delivered to the liver required for therapeutic correction of serum bilirubin. Like AAV, the liver-specific first-generation AdV also provided sustained correction in this relevant disease model. UGT1A1 mRNA expression provided per genome was comparable for both vectors. Flanking the expression cassette in AdV with AAV-ITRs (inverted terminal repeats), increased UGT1A1 mRNA expression eightfold which resulted in a significant improvement of efficacy. Compared with AAV, less AdV genomes were needed for complete correction of hyperbilirubinemia.

[Abdominal perforator flap (DIEP) and autologous latissimus dorsi in breast reconstruction. A retrospective comparative study about the first 60 cases of a same surgeon]. / Le lambeau perforant abdominal (DIEP) et grand dorsal autologue en reconstruction mammaire. Étude rétrospective comparative à propos des 60 premières patientes d'un même opérateur.

INTRODUCTION: Autologous techniques for breast reconstruction get the best cosmetic results. Aesthetic satisfaction with breast reconstruction is an important evaluation criterion. The indication is based on technical criteria (morphological, medical history) and the wishes of the patient. A rigorous evaluation of the results is necessary to assist the patients in their choice of reconstruction. METHODS: Thirty-three DIEP and 27 latissimus dorsi were involved. A satisfaction questionnaire was sent to patients to collect the aesthetic evaluation of their reconstructed breast, sequelae at the donor site of the flap as well as their overall satisfaction. Post-operative photographs of the patients were subject to aesthetical evaluation by two groups of observers. Complications were analyzed. RESULTS: The DIEP tends to get higher aesthetic satisfaction regarding the symmetry of the breasts and the volume of the reconstructed breast (P=0.05), and a better overall satisfaction (P=0.02). The uniformity of the colour of the reconstructed breast was considered superior by observers in the latissimus dorsi group (P=0.005). Donor site scar of DIEP was considered more unsightly while the latissimus dorsi was considered more painful (P=0.04) and uncomfortable, with more frequently contour abnormalities (P=0.03). We noted two total flap necrosis and three partial necrosis in the group DIEP, and two partial flap necrosis in the group latissimus dorsi. CONCLUSION: This study provides evidence that can guide the patient and the surgeon in the complex process of therapeutic decision, without exempting the latter from a careful selection of indications.

[The blood donors' haemovigilance in France]. / L'hémovigilance des donneurs de sang en France.

This work aim to present the descriptive analysis of serious adverse reactions in donors (dSAR's), which were notified in 2010 and 2011 in the French national haemovigilance database "e-FIT" (Internet secured haemovigilance reporting system). Some data, which are necessary for this analysis, also come from the regional haemovigilance coordinators' reports (RHC). The other parts of haemovigilance in the context of donation, without donors adverse reactions, such as post-donation information (PDI), adverse events occurred in the blood collection steps of the transfusion chain and epidemiology are not subject to this work analysis. This work shows that the quality of the data gradually improved since the setting up of the notification system of dSAR's. These data are particularly rich in learning lessons, but are still improving. It allows us to confirm that donor's safety, blood components quality, while preserving the blood components self-sufficiency in France, remains a priority. For these reasons, it is important to continue this haemovigilance awareness and to implement necessary actions that would be required for the protection of the donor's health and comfort during donation.

No tumour-initiating risk associated with scAAV transduction in newborn rat liver.

Delivery of recombinant adeno-associated virus (rAAV) vectors to the newborn liver is followed by a rapid loss of episomal vector copies because of hepatocyte proliferation. In selected hepatocytes, integration of rAAV genomes can lead to a sustained expression of the transgene. The safety of in vivo gene therapy with single-stranded AAV vectors has been questioned in a study reporting a high incidence of hepatocellular carcinoma, associated with provirus integration events in mice that receive an single-stranded AAV injection at birth. To investigate the tumour-initiating potential of the newly established self-complementary AAV (scAAV) vectors in the liver, groups of newborn rats received intravenous injection of a scAAV vector encoding the green fluorescent protein (GFP), or were injected with phosphate-buffered saline (PBS) or diethylnitrosamine (DEN), a well-known liver tumour initiator. The rats were fed on a diet containing 2-acetylaminofluorene, a potent liver tumour-promoting agent to accelerate the carcinogenic process. After 2 months, the animals were killed and their livers analysed. Preneoplastic nodules were identified by glutathion S-transferase-p (GSTp) staining, and GFP expression was detected by immunohistochemistry. Vector genome integration events were analysed. The numbers of GSTp-positive foci were comparable in the PBS and the scAAV-GFP groups and significantly higher in the DEN group. The proportion of GSTp-positive foci that also expressed GFP was low and in the range expected for random occurrence. No specific integration hot spots were detected by linear amplification-mediated-PCR in transduced liver. In conclusion, scAAV transduction of newborn rat liver does not trigger preneoplastic lesions suggesting an absence of liver tumourigenesis.

[Characteristics of Dupuytren's disease in women. A study of 67 cases]. / Particularités de la maladie de Dupuytren chez la femme. À propos de 67 cas.

OBJECTIVES: The aim of this study was to identify the clinical differences of the Dupuytren's disease in gender. Testosterone induces an increase of the Dupuytren's fibroblast proliferation via androgen's receptors. Testosterone rate increases during pregnancy and menopausis. We also reached a link between this factors and the clinical aspects of Dupuytren' disease in the women of our study. METHODS: This retrospective, comparative study was about all women and a randomized number of men, who underwent surgery for Dupuytren' disease between 1980 and 2010. We analysed all the epidemiologic and clinical data, the surgery procedures and the complications. Pre- and postoperative measurements of the extension lack of all the joints were performed with a manual goniometer. Disabilities of the Arm, Shoulder and Hand (DASH) questionnaire was used to evaluate the patients function. This specific data of women were reached. RESULTS: Sixty-seven women and 69 men were compared. The complex regional pain syndrome was significantly more common in women and the correction of the proximal interphalangeal joint was significantly lower in women. Recurrence rate and mean follow up were not statistically different. Mean DASH score was higher in women. We have not found any association between menopausis, pregnancy and the average age at presentation of the disease, the recurrence rate or the extension rate. CONCLUSIONS: The prognosis of the Dupuytren's disease is worse in women than in men. Other studies are necessary to reach the link between the testosterone and the clinical history of the disease in women.

[Macro-replantation of the upper limb: long-terms results and quality of life]. / Macro-replantation du membre supérieur: résultats à long terme et évaluation de la qualité de vie.

OBJECTIVES: The authors suggest standardized questionnaires of quality of life to perform a long term evaluation of patients who underwent a replantation of the proximal upper limb. METHODS: This retrospective study is about patients who underwent a replantation of the proximal upper limb between 1979 and 2011. The functional assessment was conducted using several tools including the 400 points test, the sensory tests, the Disabilities of the Arm, Shoulder and Hand questionnaire and the CHEN's classification. Some questionnaires, like the Medical Outcome Study Short Form-36 dealt with the physical and the psychological sides of the quality of life. Other questionnaires were more specific, like the Body Image Scale. RESULTS: Sixteen patients were included. The survival rate of the replanted limb was 75%. The mean follow-up was 12.7 years. We noted 20% good results with a mean DASH score to 24.5%. The quality of life was similar to the general population in most of 50% of cases. Some patients had depressive symptoms or body image troubles. CONCLUSIONS: In our eyes, the evaluation of quality of life seems essential for these patients. Survival rates, functional results, follow-up, professional activity, gender and body image troubles influence the quality of life. Besides, the physical side and the psychological side must change together to affect the overall results of the quality of life.

Insect-resistant biotech crops and their impacts on beneficial arthropods.

With a projected population of 10 billion by 2050, an immediate priority for agriculture is to achieve increased crop yields in a sustainable and cost-effective way. The concept of using a transgenic approach was realized in the mid-1990s with the commercial introduction of genetically modified (GM) crops. By 2010, the global value of the seed alone was US $11.2 billion, with commercial biotech maize, soya bean grain and cotton valued at approximately US $150 billion. In recent years, it has become evident that insect-resistant crops expressing δ-endotoxin genes from Bacillus thuringiensis have made a significant beneficial impact on global agriculture, not least in terms of pest reduction and improved quality. However, because of the potential for pest populations to evolve resistance, and owing to lack of effective control of homopteran pests, alternative strategies are being developed. Some of these are based on Bacillus spp. or other insect pathogens, while others are based on the use of plant- and animal-derived genes. However, if such approaches are to play a useful role in crop protection, it is desirable that they do not have a negative impact on beneficial organisms at higher trophic levels thus affecting the functioning of the agro-ecosystem. This widely held concern over the ecological impacts of GM crops has led to the extensive examination of the potential effects of a range of transgene proteins on non-target and beneficial insects. The findings to date with respect to both commercial and experimental GM crops expressing anti-insect genes are discussed here, with particular emphasis on insect predators and parasitoids.

Critical assessment of lifelong phenotype correction in hyperbilirubinemic Gunn rats after retroviral mediated gene transfer.

Among inherited diseases of the liver, Crigler-Najjar type 1 disease (CN-1), which results from complete deficiency in bilirubin UDP-glucuronosyltransferase activity (B-UGT1), is an attractive target for gene therapy studies. Hyperbilirubinemic Gunn rats, a model of CN-1, were injected at 2 days of age with lentiviral or oncoretroviral vectors encoding the human B-UGT1. After injection, bilirubinemia was normalized for up to 95 weeks. Bilirubin conjugates were present in the bile, demonstrating liver transduction. PCR and enzyme activity analysis confirmed gene and phenotype correction in liver. We observed that when using a strong viral promoter, a complete correction was achieved with less than 5% of B-UGT1 copy per haploid genome and after a reconstitution of 12% B-UGT1 normal activity. Liver histology remained normal throughout the experiment and tissue distribution analysis revealed preferential hepatocyte transduction after systemic delivery. Finally, no adverse immune response occurred even after induction of nonspecific liver inflammation, suggesting immune ignorance to the therapeutic protein. Our present results document the lifelong safety of gene therapy for CN-1 with retroviral vectors. They offer a better delineation of liver gene correction level required to achieve complete correction of bilirubinemia and pave the way for future clinical application of gene therapy for inherited liver disorders.

Transgenic plants for insect pest control: a forward looking scientific perspective.

One of the first successes of plant biotechnology has been the creation and commercialisation of transgenic crops exhibiting resistance to major insect pests. First generation products encompassed plants with single insecticidal Bt genes with resistance against major pests of corn and cotton. Modelling studies predicted that usefulness of these resistant plants would be short-lived, as a result of the ability of insects to develop resistance against single insecticidal gene products. However, despite such dire predictions no such collapse has taken place and the acreage of transgenic insect resistance crops has been increasing at a steady rate over the 9 years since the deployment of the first transgenic insect resistant plant. However, in order to assure durability and sustainability of resistance, novel strategies have been contemplated and are being developed. This perspective addresses a number of potentially useful strategies to assure the longevity of second and third generation insect resistant plants.

Impact of oilseed rape expressing the insecticidal serine protease inhibitor, mustard trypsin inhibitor-2 on the beneficial predator Pterostichus madidus.

Abstract Insect-resistant transgenic plants have been suggested to have deleterious effects on beneficial predators feeding on crop pests, through transmission of the transgene product by the pest to the predator. To test this hypothesis, effects of oilseed rape expressing the serine protease inhibitor, mustard trypsin inhibitor -2 (MTI-2), on the predatory ground beetle Pterostichus madidus were investigated, using diamondback moth, Plutella xylostella as the intermediary pest species. As expected, oilseed rape expressing MTI-2 had a deleterious effect on the development and survival of the pest. However, incomplete pest mortality resulted in survivors being available to predators at the next trophic level, and inhibition studies confirmed the presence of biologically active transgene product in pest larvae. Characterization of proteolytic digestive enzymes of P. madidus demonstrated that adults utilize serine proteases with trypsin-like and chymotrypsin-like specificities; the former activity was completely inhibited by MTI-2 in vitro. When P. madidus consumed prey reared on MTI-2 expressing plants over the reproductive period in their life cycle, no significant effects upon survival were observed as a result of exposure to the inhibitor. However, there was a short-term significant inhibition of weight gain in female beetles fed unlimited prey containing MTI-2, with a concomitant reduction of prey consumption. Biochemical analyses showed that the inhibitory effects of MTI-2 delivered via prey on gut proteolysis in the carabid decreased with time of exposure, possibly resulting from up-regulation of inhibitor-insensitive proteases. Of ecological significance, consumption of MTI-2 dosed prey had no detrimental effects on reproductive fitness of adult P. madidus.

Liver gene therapy: advances and hurdles.

Liver gene therapy is being developed as an alternative to orthotopic liver transplantation, which is the only effective therapy for many liver diseases. The liver has unique features that make it attractive for in vivo and ex vivo gene transfer. In vivo approach is far less invasive than ex vivo approach, although in most cases, host immune response directed against the transgene product and/or vector particles severely impairs the efficiency of gene transfer, and precludes long-term transgene expression after in vivo gene delivery. Ex vivo approach allows for an elective targeting of the hepatocytes, avoiding that the transgene be expressed in professional antigen-presenting, but is faced with the low in vitro proliferative ability of hepatocytes, and to the low in vivo liver repopulating ability of transplanted cells. In some specific situations where immune response was controlled or transplanted cells had a strong growth advantage over host hepatocytes, gene transfer resulted in long-term and complete correction of a liver genetic defect. In this review, we will outline the liver diseases that may benefit from gene therapy, the vector technology under investigation, the advances and the problems to be overcome.

Impact of oilseed rape expressing the insecticidal cysteine protease inhibitor oryzacystatin on the beneficial predator Harmonia axyridis (multicoloured Asian ladybeetle).

Insect-resistant transgenic plants have been suggested to have deleterious effects on beneficial predators through transmission of the transgene product by the pest to the predator. To test this hypothesis, effects of oilseed rape expressing the cysteine protease inhibitor oryzacystatin-1 (OC-1) on the predatory ladybird Harmonia axyridis were investigated using diamondback moth Plutella xylostella as the pest species. As expected, oilseed rape expressing OC-1 had no effects on either development or survival of the pest, which utilizes serine digestive proteases. Immunoassays confirmed accumulation of the transgene product in pest larval tissues at levels of up to 3 ng per gut. Characterization of proteolytic digestive enzymes of H. axyridis demonstrated that larvae and adults utilize cysteine and aspartic proteases; the former activity was completely inhibited by oryzacystatin in vitro. However, when H. axyridis larvae consumed prey reared on OC-1 expressing plants over their entire life cycle, no significant effects upon survival or overall development were observed. The inhibitor initially stimulated development, with a shortening of the developmental period of the second instar by 27% (P < 0.0001) accompanied by a 36% increase in weight of second instar larvae (P = 0.007). OC-1 had no detrimental effects on reproductive fitness of adult H. axyridis. Interestingly there was a significant increase in consumption of OC-1 dosed prey. The results show that prey reared on transgenic plants expressing a protein which inhibited ladybird digestive enzymes in vitro had no effects in vivo; the ladybird was able to up-regulate digestive proteases in response to the inhibitor.

Efficient retroviral gene transfer to the liver in vivo using nonpolypeptidic mitogens.

Recombinant retroviral vectors are attractive tools for achieving sustained expression of a therapeutic gene in the liver. However, cell division is required for efficient transduction with these vectors. Here we report that two widely used liver mitogens, triiodothyronin (T3) and cyproterone acetate (CPA), enable hepatocyte transduction with recombinant retroviral vectors delivered in vivo into the bloodstream. Treatment with T3 as well as CPA, alone or in combination, resulted in an increase in hepatocyte replication predominantly around the portal tract. The mitogenic activity made it possible to transduce hepatocytes in the same location. Moreover, when administered together, the two drugs synergized and the transduction level reached 5% of hepatocytes. This transduction level is compatible with clinical applications for a number of inherited liver diseases. Since these two compounds have a long history of safe clinical use, we propose that these liver mitogens may have potential for clinical application in liver-directed gene therapy.

Immune-dependent distant bystander effect after adenovirus-mediated suicide gene transfer in a rat model of liver colorectal metastasis.

Gene transfer of the herpes simplex virus thymidine kinase (HSV-tk) gene sensitizes tumor cells to the toxic effect of ganciclovir (GCV). The toxic effect of GCV extends to nontransduced surrounding cells by a metabolic process known as the bystander effect. A distant bystander effect, which involves anatomically separated tumors, has been reported in vivo. Our aim was to evaluate and characterize such distant effect in a rat model of colorectal tumors implanted in the liver using adenovirus to carry the HSV-tk gene. Two colorectal tumors were implanted in two distinct liver lobes of the liver. One of the tumor was transduced with an adenoviral vector containing HSV-tk gene. The volumes of the tumors were monitored after GCV treatment. Implication of the immune system was studied histologically and after in vivo manipulations. After GCV administration, the nontransduced distant tumor regressed partially or completely in the experimental group. Immunohistochemical analysis revealed the presence of CD8+ lymphocytes in the distant lesion. HSV-tk/GCV-induced immune response against tumors was evidenced by an adoptive transfer assay (Winn assay) and the distant bystander effect was blunted after CD8+ lymphocytes depletion. However, the survival rates for treated animals were not improved. These findings demonstrate that an immune-mediated effective distant bystander effect can be obtained after limited adenoviral-mediated transfer of the HSV-tk gene.

Critical aspects of viral vectors for gene transfer into the kidney.

Viral vectors have been used in vitro and in vivo for more than a decade, with some significant results in specific situations, e.g., when recombinant adeno-associated virus is used for the long-term transduction of skeletal muscle in coagulation factor IX-deficient patients. However, the kidney has been quite difficult to transduce with any viral vector currently available. When viral transduction occurs, it is often heterogeneous, transient, and eventually associated with immune and toxic side effects. However, recombinant adeno-associated virus and lentiviral vectors remain to be fully evaluated in the kidney; the former is small enough to be filtered through the glomerular basement membrane. This may be critical, because glomerular filtration is required for DNA complex-mediated transduction of tubular cells. An alternative to in situ renal gene transfer is secretion of a therapeutic protein from a distant site, such as skeletal muscle. Several examples provide evidence that this could be a clinically relevant approach. It also may allow accurate determination of the pathophysiologic mechanisms involved in the establishment and maintenance of experimental glomerulonephritis.

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.

In vivo retrovirus-mediated gene transfer into lamb liver.

TOPIC: Highly efficient retrovirus-mediated gene transfer into hepatocytes in vivo has been previously reported in the rat. Before considering human applications of these techniques in the treatment of inherited liver diseases, it was necessary to document its efficiency in a large animal model. Lamb was choosen because the liver was similar to human liver regarding size and anatomy. MATERIALS AND METHODS: To induce hepatocyte division which is necessary for infection with retroviral particles, animals were subjected to a left hepatectomy. Kinetics of liver regeneration were assessed on sequential liver biopsies after partial hepatectomy in order to provide an evaluation of the peak of maximal liver regeneration in a first animal group. Recombinant retroviruses encoding a reporter gene (E. coli beta galactosidase) were then perfused through the portal vein of the regenerating liver in a second animal group. RESULTS: The more intense liver regeneration occurred from one to 6 days after partial hepatectomy, with the highest thymidine kinase rate and MIB-1 antibody staining on the second day. The proportion of genetically modified lamb hepatocytes expressing the reporter gene was less than 1%, despite the use of higher titers of retroviral particles than those described in previous reports. CONCLUSION: The results obtained in rodent livers with this in vivo gene transfer methodology cannot currently be scaled up in a large ruminant model. The efficacy of vectors has to be tested in other large mammals before planning gene therapy trials for the treatment of inherited liver diseases.

In vivo retrovirus-mediated gene transfer to the liver of dogs results in transient expression and induction of a cytotoxic immune response.

Gene transfer in regenerating dog liver using high-titer recombinant retroviral vectors carrying the E. coli beta-galactosidase gene was studied. Supernatants containing amphotropic or gibbon ape pseudotyped recombinant retroviruses were infused into a peripheral vein in beagle dogs after partial hepatectomy. The kinetics of liver regeneration were determined in the animals and daily infusions were carried out for 4 or 5 days during the regeneration period. Up to 2.8% of hepatocytes were beta-galactosidase positive at the end of the procedure. However, the number of positive cells declined rapidly and few positive hepatocytes were detected after 3 weeks. PCR demonstrated the disappearance of the provirus. Histologically, inflammatory lesions were observed in the transduced livers. Finally, we demonstrated the presence of a cytotoxic T lymphocyte immune response directed against beta-galactosidase-expressing cells, which could explain the disappearance of the transgene. This work suggests that the efficiency of in vivo gene delivery using high-titer retroviral vectors directly infused into the circulation may be hampered by a cytotoxic immune response against the infected cells.