Behavioural and physiological indicators of shelter dogs' welfare: reflections on the no-kill policy on free-ranging dogs in Italy revisited on the basis of 15 years of implementation.

The Italian National Law 281 of 1991 forbids the euthanatization of free-ranging dogs, unless they have an incurable illness or are proved to be dangerous. Without neglecting the undeniable benefits of the "no-kill" policy, nevertheless it has brought about a chronic overpopulation in shelters and, as a result, higher costs of management and welfare problems since some dogs remain in the shelter for life. In 2004-2008, the Istituto Zooprofilattico Sperimentale of the Lazio and Tuscany regions carried out a survey in the Lazio Region to verify the effects of the Italian National Law 281/91 on free-ranging dog management following 15 years from its implementation. One of the aims of the study was an assessment of the welfare of dogs in a shelter sample (8 shelters out of 47 censused in the Lazio Region). 97 mixed-breed dogs were selected, their behaviour was studied and a blood sample was taken for each dog in order to determine the individual blood concentration of cortisol and the amount of oxidative damage (level of dRoms), as well as the amount of antioxidants to cope with it. Moreover, the total leukocyte count (leukogram) was accomplished. We ran general backward stepwise regression models using "level of antioxidant", "level of dRoms" and "level of serum cortisol" as dependent variables respectively. The results showed that the most important variable that improved the level of welfare of dogs consisted in having the opportunity to regularly go out of the cage for a walk, whereas other variables like gender, size of the cage (small, medium, large), being alone in the cage, and being neutered/entire, had no significant effect on the physiological indicators of welfare. Dogs that enjoyed the regular walk had a higher total antioxidant capacity, and performed a lower frequency of displacing activities and stereotyped behaviour. Moreover, oxidative stress parameters seem to be indicators well matched with behavioural indicators of stress. Thus, for the first time, markers of oxidative status are utilised for the welfare evaluation in the domestic dog. Furthermore, the results of this paper give some suggestion about how small steps can help to improve shelters and, furthermore, this paper intends to solicit the debate on the no-kill policy.

Role of the ventral striatum in developing anorexia nervosa.

Functional imaging data in adult patients with anorexia nervosa (AN) support a dysfunctional signal in the ventral striatum as neural signature of AN. In the present study, development of this signal was investigated with the prediction that a characteristic pattern of ventral-striatal signalling will be shown in response to cues associated with food restriction that reflects the evolvement of starvation dependence over time. The signal was assessed in adolescent patients with AN, whose duration of illness was about five times shorter relative to the adult sample. During functional magnetic resonance imaging subjects were required to estimate weights of body images (underweight, normal weight, overweight) and to process each stimulus in a self-referring way. Relative to age-matched, young healthy controls, underweight stimuli were already associated with greater activity of the ventral striatum, and processing of normal-weight stimuli elicited already reduced signalling. Subjective preferences showed exactly the same pattern of results. Relative to adult AN, the present data reveal a developing dysfunctional signal that, if untreated, will essentially contribute to the maintenance of AN. We discuss putative mechanisms that may play a crucial role in the development of AN, and also deduce new hypotheses about the involvement of the midbrain dopamine system, of which illness-related alterations may contribute to the development of AN.

Biotic and abiotic factors influencing distribution and abundance of Culicoides obsoletus group (Diptera: Ceratopogonidae) in central Italy.

In the framework of a bluetongue surveillance program including clinical, serological, and entomological activities, Culicoides biting midges were light trapped weekly in two regions of central Italy, Lazio and Tuscany. In the period January 2002 through December 2005, 3,944 collections were carried out in 189 trap sites distributed in all the provinces of the two regions. Abundance data of C. obsoletus group were analyzed in relation to trap site altitude, distance from the sea, land use, and number of farmed animals. Species seasonality and overall temporal trend were also described. C. obsoletus was distributed over the whole study area, almost in all trapping sites and with high abundances. The species group was dominant among all captured Culicoides, with higher abundances recorded inland and in areas where land cover was partially or completely natural-wooded. Adults on the wing were caught all year round, with peaks in May-June and middle October. The observed trend through years recorded a peak during autumn 2002, in concomitance with a local epidemic of bluetongue.

Mek1/2 gene dosage determines tissue response to oncogenic Ras signaling in the skin.

Ras genes are commonly mutated in human cancers of the skin and other tissues. Oncogenic Ras signals through multiple effector pathways, including the Erk1/2 mitogen-activated protein kinase (MAPK), phosphatidylinositol-3 kinase (PI3K) and the Ral guanine nucleotide exchange factor (RalGEF) cascades. In epidermis, the activation of oncogenic Ras induces hyperplasia and inhibits differentiation, features characteristic of squamous cell carcinoma. The downstream effector pathways required for oncogenic Ras effects in epidermis, however, are undefined. In this study, we investigated the direct contribution of Mek1 and Mek2 MAPKKs to oncogenic Ras signaling. The response of murine epidermis to conditionally active oncogenic Ras was unimpaired by deletion of either Mek1 or Mek2 MAPKKs individually. In contrast, Ras effects were entirely abolished by combined deletion of all Mek1/2 alleles, whereas epidermis retaining only one allele of either Mek1 or Mek2 showed intermediate responsiveness. Thus, the effects of oncogenic Ras on proliferation and differentiation in skin show a gene dosage-dependent requirement for the Erk1/2 MAPK cascade at the level of Mek1/2 MAPKKs.

A case of an infant with flail tricuspid valve due to spontaneous papillary muscle rupture: was neonatal lupus the culprit?

A 3-month-old infant presented in extremis with a flail tricuspid valve. The authors theorized that the genesis of her papillary muscle rupture was perinatal ischemia compounded by worsening pulmonary valvular stenosis leading to excessive fiber tension. Her underlying diagnosis of autoimmune-mediated heart block with endocardial fibroelastosis and prenatal glucocorticoid steroid treatment represents potentiating factors.

Active circulation of bluetongue vaccine virus serotype-2 among unvaccinated cattle in central Italy.

Several seroconversions occurring in 2002 among sentinel cattle during the bluetongue-vaccination campaign in Lazio and Tuscany (central Italy) led to the suspicion of vaccine-virus circulation. Therefore in 2003, 17 seroconverting sentinel herds were investigated for the characteristics of the virus involved. From these farms, 91 unvaccinated animals and 57 Culicoides pools were tested for the presence of the bluetongue vaccine virus (serotype-2) or other strains. The presence of vaccine virus serotype-2 was confirmed by PCR followed by restriction analysis in the whole blood of 17 unvaccinated sentinel cattle and 12 pools of Culicoides imicola or C. obsoletus. Of the 17 herds, five were positive only for vaccine virus serotype-2, four were positive for other strains and two for both the vaccine and other strains; the remaining premises were virologicaly negative. The vaccine virus serotype-2 also was detected in areas not included in the vaccination campaign.

Association between the 2001-2003 bluetongue epidemic in Lazio and Tuscany (central Italy) and distribution and abundance of Culicoides imicola and C. obsoletus vectors.

During the epidemic of bluetongue (BT) in Lazio and Tuscany between 2001 and 2003, the distribution pattern of Culicoides imicola did not always correspond either geographically or seasonally, with virus circulation. Culicoides obsoletus was observed to be abundant, ubiquitous and active throughout the year. The geographical and seasonal distribution of BT virus (BTV), C. imicola and C. obsoletus was compared. The territory of the two regions was divided into 30 cells each measuring 1 600 km(2). The presence of C. obsoletus was recorded in every cell, while C. imicola was detected in 18 of the 30 cells, but was absent in 6 of the 21 cells that indicated the presence of BTV. The occurrence of seroconversions appeared to be positively correlated with maximum C. obsoletus catches. Seroconversions were recorded throughout the year, even when C. imicola was not active, whereas C. obsoletus was detected during the entire period. The occurrence of BTV circulation in areas and periods where C. imicola was absent, and the abundant and constant presence of adult C. obsoletus in all the cells, suggest the active role of the latter species in BTV circulation in central Italy.

Geographical and seasonal distribution of the bluetongue virus vector, Culicoides imicola, in central Italy.

Following the first incursion of bluetongue virus (BTV) into Italy, the geographical and seasonal distribution of the biting midge Culicoides imicola Kieffer (Diptera: Ceratopogonidae), the main vector of BTV and African horse sickness virus, was investigated in two regions of central Italy (Lazio and Tuscany). Surveillance of Culicoides was carried out between July 2001 and December 2002 using light traps: 1917 collections were made in 381 trap sites, well distributed across both regions. During the survey, bluetongue outbreaks were recorded in both regions. Culicoides imicola was found in 89 (23%) trap sites, distributed fairly continuously along the whole western coastline, between 41.2697 degrees N and 44.05724 degrees N. It was found only occasionally inland and usually in low abundance, with catches of more than 1000 specimens per night found in only two sample sites and 74% of catches numbering fewer than 10 specimens. Adults were caught from March to mid December, with peaks ranging from the end of August to mid November. The coastal distribution and the presence of only few sites with year-round records of adult vectors suggests that colonization may have occurred recently, by passive wind-dispersal from external source areas (Sardinia and Corsica). Alternatively, the species may occur in established, previously undetected, autochthonous populations that are limited from extension inland and northern-ward within Lazio and Tuscany by cool winter temperatures.

Effects of human tissue plasminogen gene transfer on allograft coronary atherosclerosis.

BACKGROUND: Transplant coronary atherosclerosis is a major limiting factor to successful long-term cardiac transplantation. The depletion of tissue plasminogen activator (tPA) in the arteriolar smooth muscle cells has been associated with a higher incidence of accelerated graft atherosclerosis. In vivo overexpression of tPA may inhibit accelerated graft atherosclerosis and improve the long-term results of heart transplantation. We evaluated the feasibility, distribution, and effects of intracoronary transfer of the human tPA (htPA) gene in a rabbit heterotopic cardiac transplant model, using a novel cationic liposome compound designed for improved delivery to vascular endothelium. METHODS: Human tPA cDNA under the control of the SV40 promoter (100 microg) was complexed with the novel cationic liposome (+/-)-N-(3-aminopropyl)-N,N-dimethyl-2,3-bis(dodecyloxy)-1-propanaminium bromide (GAP: DLRIE) (50 microg), and delivered ex vivo to the donor heart by slow intracoronary infusion. Control hearts received an "empty" liposome preparation. Grafts were then implanted into recipient rabbits in the heterotopic cervical position. For the analysis of gene expression, beating donor hearts were collected at 4 days. To examine the effects of htPA expression on graft atherosclerosis, animals received a 0.5% cholesterol diet for 30 days posttransplant, as well as 10 mg/kg cyclosporine A daily. Beating hearts were collected at 30 days posttransplant and analyzed for the development of transplant atherosclerosis by image analysis. RESULTS: Northern blot analysis for the htPA messenger RNA (mRNA) transcripts showed significantly higher counts in hearts receiving the htPA gene as compared to controls. The distribution of these transcripts favored the left ventricle (LV) and septal regions over the right ventricle (RV). Scintillation analysis of specimens stained by immunoflourescence showed expression of htPA throughout the perivascular myocardium that was significantly higher in grafts transduced with the htPA gene than in control or native hearts. Expression in the vascular wall was also significantly enhanced. Scintillation counts per x 200 field were 262 +/- 145 in htPA-transduced hearts and 20 +/- 27 in controls (p = 0.001), and mean luminescence was 83.7 +/- 12.5 in htPA-transduced hearts and 62.9 +/- 12.8 in controls (p = 0.01). Intimal hyperplasia was assessed by mean percent luminal stenosis in small- and medium-sized arteries and was 31.12 +/- 23.5% in htPA-transduced hearts and 86.59 +/- 17.5% in control hearts (p < 0.0001). These results demonstrate that expression of the htPA gene can be induced by ex vivo intracoronary gene transfer at the time of allograft preservation. Liposome-mediated delivery of the htPA gene at the time of transplantation results in significant early transgene expression, and significantly inhibits the development of graft coronary atherosclerosis.

PAX3 is expressed in human melanomas and contributes to tumor cell survival.

Deregulated expression of the transcription factor PAX3 was observed previously in several tumors like rhabdomyosarcoma and Ewing's sarcoma. Because PAX3 expression is also found in pluripotent neural crest cells, we investigated whether melanomas, tumors derived mostly from cutaneous intraepidermal melanocytes, might show deregulated PAX3 expression. Using a specific and sensitive reverse transcription-PCR, we detected PAX3 mRNA in 77% (27 of 35) of primary cultured melanomas. These results could be confirmed by direct in situ hybridization on the corresponding tissue sections where PAX3 expression was unambiguously confined to tumor cells and not detected in surrounding normal tissue, normal skin sections, or sections of benign lesions. Furthermore, down-regulation of PAX3 expression achieved through a specific antisense oligonucleotide-based treatment resulted in > 70% of dead cells specifically in PAX3-positive melanomas. Annexin V staining confirmed that primary melanoma cells underwent apoptosis after treatment These experiments suggest that in situ hybridization of PAX3 on paraffin-embedded tissue may represent a novel means to identify melanoma cell lesions, which appear to become dependent on expression of this deregulated transcription factor.

DRAL is a p53-responsive gene whose four and a half LIM domain protein product induces apoptosis.

DRAL is a four and a half LIM domain protein identified because of its differential expression between normal human myoblasts and the malignant counterparts, rhabdomyosarcoma cells. In the current study, we demonstrate that transcription of the DRAL gene can be stimulated by p53, since transient expression of functional p53 in rhabdomyosarcoma cells as well as stimulation of endogenous p53 by ionizing radiation in wild-type cells enhances DRAL mRNA levels. In support of these observations, five potential p53 target sites could be identified in the promoter region of the human DRAL gene. To obtain insight into the possible functions of DRAL, ectopic expression experiments were performed. Interestingly, DRAL expression efficiently triggered apoptosis in three cell lines of different origin to the extent that no cells could be generated that stably overexpressed this protein. However, transient transfection experiments as well as immunofluorescence staining of the endogenous protein allowed for the localization of DRAL in different cellular compartments, namely cytoplasm, nucleus, focal contacts, as well as Z-discs and to a lesser extent the M-bands in cardiac myofibrils. These data suggest that downregulation of DRAL might be involved in tumor development. Furthermore, DRAL expression might be important for heart function.

Ectopic expression of PA2.26 antigen in epidermal keratinocytes leads to destabilization of adherens junctions and malignant progression.

PA2.26 antigen is a small mucin-type transmembrane glycoprotein induced in mouse epidermal keratinocytes during carcinogenesis. It is located at plasma membrane projections, such as microvilli and ruffles, where it interacts with the actin cytoskeleton. Previous studies revealed that ectopic expression of PA2.26 in epidermal MCA3D keratinocytes induces cell surface extensions and increased motility. Here, we show that PA2.26-expressing MCA3D (3D2.26) cell transfectants undergo a phenotypic conversion linked to the acquisition of malignant characteristics. The 3D2.26 cells down-regulate basal keratin K14 and up-regulate vimentin and keratin K8 expression. Immunofluorescence analysis in 3D2.26 cell cultures showed loss of cortical actin filaments and destabilization of adherens junctions mediated by E- and P-cadherin, although both cadherin mRNAs were expressed in the transfectants. When the cadherin protein levels were analyzed in Western blots, no P-cadherin protein or smaller polypeptide E-cadherin forms were detected, suggesting that E- and P-cadherin synthesized in 3D2.26 cells was unstable and proteolytically degraded. Transplantation of 3D2.26 cells into athymic nude mice induced tumors, whereas MCA3D cells and control (3DN) transfectants were not tumorigenic after 72 days postinjection. The phenotype of the tumors was undifferentiated, with mixed regions exhibiting a glandular differentiation pattern in which the presence of numerous surface microvilli was observed at the ultrastructural level. Interestingly, PA2.26 antigen was highly expressed in these microvillous cell surfaces. Tumor cells were vimentin- and K8-positive and showed an aberrant pattern of E-cadherin protein expression in which large cytoplasmic aggregates were found close to the nucleus. Infiltration of tumor cells into lymphatic vessels and the presence of frequent regional lymph node metastases were also observed in the tumors. These results indicate that expression of PA2.26 antigen in premalignant keratinocytes induces a fully transformed and metastatic phenotype, and they suggest an involvement of PA2.26 in malignant progression.

Inv(11)(p13p15) and myf-3(MyoD1) in a malignant extrarenal rhabdoid tumor of a premature newborn.

We present a premature newborn of 32 wk of gestation with a congenital malignant extrarenal rhabdoid tumor (MERT) on the right shoulder with generalized metastases. Standard histologic, immunohistochemical, molecular and cytogenetic methods were used in the evaluation of diagnostic material. Biopsy of a skin lesion showed the histologic features of a malignant rhabdoid tumor. Cytogenetic analysis of the tumor cells revealed an inv(11)(p13p15) and additionally, an increased expression of myf-3 (myogenic determination factor, MyoD1) and PAX3 was detected. These results suggest an origin of the neoplasm derived from a pluripotent cell with the potential of myogenic differentiation. Tumor suppressor genes located on chromosome 11p13 and 11p15 may play an important role for malignant rhabdoid tumor development and progression.

Molecular features of a human rhabdomyosarcoma cell line with spontaneous metastatic progression.

A novel human cell line was established from a primary botryoid rhabdomyosarcoma. Reverse transcription polymerase chain reaction investigations of this cell line, called RUCH-2, demonstrated expression of the regulatory factors PAX3, Myf3 and Myf5. After 3.5 months in culture, cells underwent a crisis after which Myf3 and Myf5 could no longer be detected, whereas PAX3 expression remained constant over the entire period. Karyotype analysis revealed breakpoints in regions similar to previously described alterations in primary rhabdomyosarcoma tumour samples. Interestingly, cells progressed to a metastatic phenotype, as observed by enhanced invasiveness in vitro and tumour growth in nude mice in vivo. On the molecular level, microarray analysis before and after progression identified extensive changes in the composition of the extracellular matrix. As expected, down-regulation of tissue inhibitors of metalloproteinases and up-regulation of matrix metalloproteinases were observed. Extensive down-regulation of several death receptors of the tumour necrosis factor family suggests that these cells might have an altered response to appropriate apoptotic stimuli. The RUCH-2 cell line represents a cellular model to study multistep tumorigenesis in human rhabdomyosarcoma, allowing molecular comparison of tumorigenic versus metastatic cancer cells.

Identification of PA2.26 antigen as a novel cell-surface mucin-type glycoprotein that induces plasma membrane extensions and increased motility in keratinocytes.

PA2.26 antigen was identified as a cell-surface protein induced in epidermal carcinogenesis and skin remodeling processes. PA2.26 is expressed in carcinoma cell lines and cultured fibroblasts but absent in nontumorigenic keratinocytes. In tissues, PA2.26 is present in epithelial cells of the choroid plexus, ependyma, glomerulus and alveolus, in mesothelial cells, and in endothelia of lymphatic vessels. Biochemical characterization of PA2.26 protein and sequence analysis of the isolated cDNA demonstrate that PA2.26 antigen is a mucin-like transmembrane glycoprotein. Confocal and immunoelectron microscopy analysis in cultured cells reveal that PA2. 26 is concentrated in actin-rich microvilli and plasma membrane projections, such as filopodia, lamellipodia and ruffles, where it colocalizes with members of the ERM (ezrin, radixin, moesin) family protein. Ezrin and moesin, but not radixin, can be coimmunoprecipitated together with PA2.26 from cell lysates. Ectopic expression of PA2.26 in immortalized, nontumorigenic, keratinocytes induces an epithelial-fibroblastoid morphological conversion with increased plasma membrane extensions, concomitantly to a major reorganization of the actin cytoskeleton, redistribution of ezrin to cell-surface projections, and enhanced motility. These findings suggest an involvement of PA2.26 in cell migration.

Interval or permanent nonoperative management of acute type A aortic dissection.

HYPOTHESIS: Selected patients with acute type A (ascending) aortic dissection who are treated with delayed operation or nonoperative therapy may have better early and short-term outcomes than was previously expected. DESIGN AND SETTING: Retrospective cohort at a university hospital. SUBJECTS: Data on 75 patients with acute or chronic type A aortic dissection treated at one institution from January 1, 1985, to November 30, 1997, were analyzed. Of these 75 patients, 34 (21 male and 13 female, with a mean age of 65.5 years) did not undergo initial operative treatment, and 15 (10 male and 5 female, with a mean age of 72.6 years) never underwent surgery. For the 19 patients who underwent delayed surgery, the mean period between aortic dissection and intervention was 11.4+/-4.83 days. The follow-up period ranged from 0.27 to 149 months, with a mean of 20.2 months. MAIN OUTCOME MEASURES: Vascular complications, hospital mortality, and early survival. RESULTS: Reasons for interval delay in surgical treatment included initial misdiagnosis or delay in diagnosis (13 [68%] of 19), need to address significant comorbidity (4 [21%] of 19), and initial refusal of operative intervention (2 [11%] of 19). For the 15 patients treated entirely by medical therapy, reasons for electing nonoperative management included extensive comorbidity (5 [33%] of 15), refusal of surgical intervention (6 [40%] of 15), and misdiagnosis or long delay in diagnosis (4 [27%] of 15). Of the 34 patients, 15 (44%) presented with moderate or severe aortic insufficiency, 5 (14%) had evidence of pericardial effusion, 6 (21%) had evidence of concomitant coronary ischemia on electrocardiogram, and 8 (24%) had extension of the dissection into the descending aorta. Four patients (11.8%) died while in the hospital. Of the 34 patients, 30 (88%) who underwent either delayed or no surgery received aggressive medical treatment (beta-adrenergic blocking agents and afterload-reducing agents) and were discharged from the hospital. All patients who were operative candidates in the interval treatment group survived to reach definitive operation. There was no statistically significant difference in short-term survival between the group of patients undergoing delayed surgery or medical treatment only and the group of 41 patients undergoing early operation (P = .42). CONCLUSIONS: Immediate surgical therapy is still recommended for acceptable operative candidates with acute type A aortic dissection who seek immediate treatment. However, this study permits the following 2 conclusions: (1) patients with type A aortic dissection who are referred or whose conditions are diagnosed several days after presentation have survived the early dangerous period and can safely undergo surgery semielectively (rather than emergently); and (2) selected patients who are not considered operative candidates and who survive the initial type A aortic dissection without complication may be treated with aggressive medical therapy and achieve acceptable early and short-term outcomes, which is better than previously expected.

Multiple left heart obstructions (Shone's anomaly) with mitral valve involvement: long-term surgical outcome.

BACKGROUND: The outcome of children with multilevel left heart obstructions (Shone's anomaly) is generally poor. Literature is scarce, consisting mainly of case reports. The mitral disease may be the predominant factor affecting outcome. METHODS: Surgical results in 19 consecutive patients are presented, with a median follow-up of 8 years. Mitral stenosis was present in all, with parachute deformity in 12 patients. Supramitral rings were found in 9 patients. Other features included subaortic stenosis (15 patients), valvar aortic stenosis (9), bicuspid aortic valve (16), and coarctation (13 patients). The patients underwent 46 surgical procedures, including 18 mitral operations (9 replacements, 9 repairs). RESULTS: There were three in-hospital (16%) and two late (10.5%) deaths. Of the 5 nonsurvivors, 4 patients (80%) had predominant mitral disease and moderate to severe pulmonary hypertension, versus 4 (28.5%) and 5 (36%) survivors, respectively (p = not significant). Valve repair was the final procedure in 9 survivors. The other 5 patients had repeated valve replacements (1), aortoventriculoplasty with valve replacements (2), or no mitral operation (2). Freedom from mitral reoperation was 78% (7 of 9 patients) after repair procedures and 43% (3 of 7 patients) after replacement. At follow-up, 10 patients (71.4%) are in New York Heart Association functional class I and the other 4 in class II and III. Six (43%) await reoperation due to recurrent aortic (4) or subaortic (1) stenosis and recoarctation (2). Echocardiography reveals mild mitral stenosis or regurgitation in 3 patients after repair (33%). Four are considered free of residual disease (21% of all). CONCLUSIONS: Late outcome in Shone's anomaly seems to correlate with the predominance of mitral valve involvement and the degree of pulmonary hypertension. Valve repair is indicated whenever feasible and should be considered before the occurrence of pulmonary hypertension.

Induction of PA2.26, a cell-surface antigen expressed by active fibroblasts, in mouse epidermal keratinocytes during carcinogenesis.

The monoclonal antibody PA2.26, produced against mouse epidermal keratinocytes transformed with 7,12-dimethylbenz[a]anthracene (DMBA), recognizes a 43- to 47-kDa cell-surface protein that was absent from non-tumorigenic epidermal keratinocytes but present in transformed epidermal cell lines as well as cultured normal fibroblasts. In vivo, the antigen was absent from normal epidermis but induced in basal-like epidermal keratinocytes and dermal fibroblasts during tissue regeneration after wounding and treatment with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA). The PA2.26 protein was also expressed in basal-like cells of differentiated papillomas and carcinomas generated in mice treated with DMBA and TPA. In addition, the antigen was abundantly synthesized by stromal cells of the tumors. These results suggest that PA2.26 antigen is involved in reactive processes during skin remodeling and carcinogenesis.

The alternate recipient list for heart transplantation: does it work?

BACKGROUND: One quarter of patients awaiting heart transplantation die while on the waiting list. This is largely due to the shortage of donor organs. The alternate recipient list was created to establish a means by which patients who would otherwise be turned down for heart transplantation solely because of age over 65 or a need for a third heart transplantation can receive organs considered marginal that may otherwise be wasted. The hope is that these patients may achieve improved survival with these substandard hearts than they would achieve with medical therapy alone. METHODS: Twenty-two patients ages 47 to 71 years (mean 66.7 years) were listed on the alternate recipient list at the University of California at Los Angeles Medical Center from 1991 to 1996. Seventeen patients underwent heart transplantation from the alternate waiting list. The outcome of this group was compared with the outcome of a contemporaneous group of 266 patients ages 18 to 66 years (mean age 52.1 years) from the standard heart transplantation waiting list. RESULTS: The early mortality rate for the patients in the alternate group was 11.8% (2/ 17). Actuarial survival from time of orthotopic heart transplantation at 6 months and 1 year was the same 74.5% at a mean follow-up was 13.4 months. In comparison, the early mortality rate for the patients on the standard list was 5.6% (15/266), and actuarial survival at 6 months and 1 year was 86.8% and 83.1%, respectively (mean follow-up was 30 months). There was no significant difference in early mortality rate or actuarial survival between the two groups. CONCLUSION: The alternate recipient list for heart transplantation is a valid and ethical option for patients who would otherwise be denied heart transplantation. It provides these patients with similar early and medium-term outcomes in comparison to patients on the standard list, and organs that may otherwise be wasted are used.

Intracoronary gene transfer of immunosuppressive cytokines to cardiac allografts: method and efficacy of adenovirus-mediated transduction.

OBJECTIVE: Allograft-targeted immunosuppressive gene therapy may inhibit recipient immune activation and provide an alternative to systemic immunosuppression. We studied the optimal technique and efficacy of intracoronary gene transfer of viral interleukin-10 and human transforming growth factor-beta 1 in a rabbit model of heterotopic heart transplantation. METHODS: Replication-defective adenoviral vectors were constructed, expressing viral interleukin-10 (AdSvIL10) or transforming growth factor-beta 1 (AdCMVTGF-beta 1). Intracoronary delivery of vectors was accomplished ex vivo by either bolus injection or slow infusion. The allografts were implanted heterotopically in recipient rabbits and collected 4 days after the operation. Vector dose was 4 x 10(9) to 6 x 10(10) pfu/gm of donor heart. Transfer was confirmed by DNA amplification for both genes. Gene product expression in tissue was quantified by immunoassay and visualized by immunohistochemical staining. RESULTS: Allograft viral uptake was only 9.9% +/- 2.4% with bolus injection, but increased to 80.5% +/- 6.8% at 1 ml/min infusion rate (p = 5 x 10(-14)). Uptake ratio was not affected by vector quantity or slower infusion rates. Transforming growth factor-beta 1 was consistently detected in allografts infected with AdCMVTGF-beta 1, but not with control adenovirus or AdSvIL10. Expression was proportional to infused vector quantity and reached 10 ng/gm of allograft at infused 10(10) pfu/gm. Transforming growth factor-beta 1 was also detected in recipient's serum at less than 1 ng/ml. Viral interleukin-10 was detected in minor amounts only (< 1 ng/gm) in allografts infected with AdvIL10 up to 5 x 10(10) pfu/gm. Nevertheless, it was detected in recipient serum at concentrations up to 0.4 ng/ml. CONCLUSIONS: Intracoronary gene transfer of immunosuppressive cytokines to cardiac allografts during cold preservation is feasible. Slow infusion is superior to bolus injection. In vivo effects on allograft rejection remain to be determined.