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A Shaked

Publications and source records attributed to A Shaked.

At least 73 records · Page 4Linked to original sources
PubMed1995

Adenovirus-mediated p53 gene delivery inhibits 9L glioma growth in rats.

Adenoviral vectors have recently been shown to effectively deliver genes into a variety of tissues. Since these vectors have some advantages over the more extensively investigated retroviruses, we studied the effect of two replication-defective adenovectors bearing human wild type tumor suppressor gene p53 (Adp53) and Escherichia coli beta-galactosidase gene (AdLacZ) on 9L glioma cells. Successful in vitro gene transfer was shown by DNA polymerase chain reaction (PCR), and expression was confirmed by reverse transcriptase RNA PCR and Western blot analyses. Transduction of 9L cells with the Adp53 inhibited cell growth and induced phenotypic changes consistent with cell death at low titers, while AdLacZ caused cytopathic changes only at high titers. Stereotactic injection of AdLacZ (10(7) plaque forming units) into tumor bed stained 25 to 30% of tumor cells at the site of vector delivery. Injection of Adp53 (10(7) plaque forming units), but not AdLacZ (controls), into established 4-day old 9L glioma brain tumors decreased tumor volume by 40% after 14 days. As a step toward gene therapy of brain tumors using replication-defective adenoviruses, these data support the use of tumor suppressor gene transfer for in vivo treatment of whole animal brain tumor models.

Adenoviridae↗
PubMed1995

Adjuvant chemotherapy improves survival after liver transplantation for hepatocellular carcinoma.

OBJECTIVE: The aim of this study was to evaluate the effect of postoperative adjuvant chemotherapy on the recurrence rate and survival of patients after orthotopic liver transplantation (OLT) for hepatocellular carcinoma (HCC). SUMMARY BACKGROUND DATA: Historically, liver transplantation for HCC has yielded poor long-term survival. Multimodality therapy has been initiated in an effort to improve survival statistics. METHODS: Twenty-five patients were placed on 6 months of intravenous fluorouracil, doxorubicin, and cisplatin after OLT. Risk factors, recurrence rates, and survival rates were analyzed and compared with historic controls. RESULTS: Overall long-term survival in the protocol patients was 46% at 3 years, improved over our historic controls of 5.8% at 3 years (p = 0.0001). Overall recurrence rate was 20% (n = 4). Possible risk factors, such as tumor size, vascular invasion, multifocality, capsular invasion, and tumor differentiation, were not found to be significantly predictive of survival. Three patients with long-term, disease-free survival had tumors > 5 cm. Side effects from chemotherapy were common, but rarely severe. CONCLUSIONS: This study suggests that adjuvant chemotherapy after transplantation for HCC can provide long-term cure and may improve survival, even in patients with stage III and IV disease.

Actuarial Analysis↗
PubMed1995

Evaluation, operative management, and outcome after liver transplantation in children with biliary atresia and situs inversus.

INTRODUCTION: Biliary atresia, a common indication for liver transplantation, can be associated with situs inversus. Our experience with liver transplantation in children (n = 6) was reviewed retrospectively. PATIENTS AND METHODS: Preoperative duplex sonography, computerized tomography, and visceral angiography were obtained. Vascular anomalies identified included preduodenal portal vein (6/6), interrupted inferior vena cava (5/6), and aberrant hepatic artery (4/6). RESULTS: The liver graft was placed in a midline position. Venous continuity was achieved by donor suprahepatic inferior vena cava to recipient hepatic cloaca and direct end-to-end portal anastomosis. The donor infrahepatic inferior vena cava was oversewn. Arterial continuity was restored using either a direct branch-patch anastomosis (3/6) or a supraceliac aortic interposition graft (3/6). In retrospect, preoperative diagnostic work-up was noncontributory and outcome was not complicated by pre-existing situs inversus. CONCLUSION: Situs inversus in liver recipients requires operative technical modifications, but does not change outcome. Furthermore, extensive preoperative work-up should be avoided.

Biliary Atresia↗
PubMed1994

The correlation of intragraft cytokine expression with rejection in rat small intestine transplantation.

Rejection continues to be a major cause of graft loss in small intestine transplantation (SIT). We have studied, by semiquantitative reverse transcriptase PCR (rtPCR), the intragraft expression of cytokines relevant to rejection in a rat model. Heterotopic SIT grafts were performed from Lewis x Brown Norway F1 donors into Lewis recipients. The isograft control was Lewis into Lewis. Five animals in each isograft and allograft group were sacrificed on POD 3, 5, 7, 8, 9, 10, 12, and 14. mRNA was isolated from portions of the terminal ileum and rtPCR performed to amplify message for interleukin-2 (IL-2), IL-2 receptor (IL-2R), interleukin-6 (IL-6), tumor necrosis factor alpha (TNF-alpha), and interferon gamma (IFN-gamma). Semiquantitative analysis was performed using 32P radionuclide incorporation and scintillation counting. The results were expressed as percent activity compared with beta-actin. Histologic correlation with cytokine expression was made. On POD 3 after SIT there was no evidence of rejection by histology and all cytokines studied showed no difference between the isograft and the allograft. On POD 5 the first evidence of mild rejection was seen on histology and IL-6, IFN-gamma, TNF-alpha showed a significant up regulation in the allograft that persisted through POD 14. mRNA for IL-2 was not significantly upregulated until POD 7 and persisted until POD 14. IL-2R was constitutively expressed in both isograft and allograft and was not a reliable predictor of rejection. Histologic rejection was moderately severe by POD 7 and severe between POD 8 and 14 correlating with the increasing expression of IL-6, IFN-gamma, and TNF-alpha. In summary, we have shown that increasing expression of mRNA for IL-6, IFN-gamma, and TNF-alpha not only correlated with severity of rejection but that upregulation began early when histologic evidence of rejection first occurred.

Animals↗
PubMed1994

Risk factors for liver transplantation across the ABO barrier.

A retrospective study was performed to identify risk factors affecting the fate of liver allografts transplanted across the ABO barrier. Over a 5-year period, 30 patients received 31 emergency OLTs with ABO-incompatible allografts. One-year graft and patient survival rates were 37.1% and 48.4%, respectively. This group was compared with 199 ABO-compatible OLTs performed under similar emergency conditions. Significantly higher incidence of rejection, hepatic artery thrombosis, and biliary stricture were found in ABO-incompatible grafts. The graft or patient survival of patients with O blood group or fulminant hepatic failure was not affected by ABO incompatibility. In contrast, the 1-year survival of recipients with non-O blood group was only 13.3% and the 1-year survival of those undergoing retransplantation was only 27.7% after ABO-incompatible OLT. A multivariate analysis using the Cox proportional hazard model identified non-O blood group (relative risk 3.43) and retransplantation (relative risk 2.46) as the most important risk factors for patient survival after ABO-incompatible OLT. We conclude that the use of ABO-incompatible grafts is justified in emergency cases when no other donor is available, but should be proscribed in patients with non-O blood group. The value of ABO-incompatible grafts in retransplantation is questionable and requires further investigation.

ABO Blood-Group System↗
PubMed1994

Adenovirus-mediated gene transfer in the transplant setting. I. Conditions for expression of transferred genes in cold-preserved hepatocytes.

Transplantation of genetically modified hepatocytes has been suggested as a therapeutic modality for impaired hepatocellular function. This study examined adenoviral-mediated gene transfer to isolated hepatocytes, under conditions mimicking clinical transplant preservation. Isolated rat hepatocytes were infected using replication-defective adenoviral vectors with an expression cassette containing the beta-galactosidase gene driven by a CMV promoter. Hepatocytes were infected in suspension immediately after isolation, then either cultured or transplanted immediately into a syngeneic host. Gene transfer efficiency was assessed by histochemical staining and FACS analysis for the gene product. The presence of viral DNA and mRNA, as well as viral-derived protein production, were assayed. Efficiency of gene transfer was examined as a function of several preservation conditions. Infection efficiency was best in cells preserved in UW solution, correlated directly with virus:hepatocyte ratio and with length of exposure to virus. Successful infection resulted in significant viral-derived protein production, persisting for at least two weeks in culture. These results demonstrate the versatility of adenoviral vectors in accomplishing rapid and efficient gene transfer into nondividing hepatocytes during cold preservation. Such genetically modified hepatocytes have potential use for immediate transplantation, without the need for further manipulation.

Adenoviridae↗
PubMed1994

Adenovirus-mediated gene transfer in the transplant setting. II. Successful expression of transferred cDNA in syngeneic liver grafts.

These experiments establish a model for gene transfer to transplanted liver grafts ex vivo using adenoviral vectors. Rat liver grafts (n = 8) were harvested, and preserved in UW or lactated Ringer's. The grafts were infected ex-vivo via portal vein perfusion with replication-defective Ad vectors encoding the beta-galactosidase (beta-gal) gene diluted in UW solution. The infected grafts were stored at 4 degrees C for 1 hr, then transplanted into syngeneic hosts. Liver biopsies were taken at 1, 7, and 15 days after transplantation. Infection rate was assessed by histochemical staining for beta-gal. Liver DNA and RNA were assayed for the beta-gal sequences, and recombinant protein production measured at 24 hr and 7 days after transplantation. Under conditions mimicking liver graft cold storage, efficient gene transfer was achieved with an infection rate of 10-15%, as assessed by X-gal staining. Viral DNA and RNA presence in the graft was confirmed; gene expression with protein production were verified by western blots and a functional protein assay. All studies were negative in control livers. Gene expression persisted for at least 2 weeks after transplantation. We conclude that efficient adenovirus-mediated gene insertion and expression of gene products can be accomplished in whole-liver grafts under hypothermic preservation conditions currently used in clinical transplantation.

Adenoviridae↗
PubMed1994

Intragraft expression of messenger RNA for interleukin-6 and tumor necrosis factor-alpha is a predictor of rat small intestine transplant rejection.

Detection of rejection after small intestine transplantation (SIT) is difficult, relying largely on histopathology. The purpose of this study was to determine if the intragraft expression of messenger RNA (mRNA) for interleukin-2 receptor (IL-2R), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF) correlated with rejection in a unidirectional, heterotopic rat SIT model. Graft samples were obtained on postoperative day (POD) 3, 5, 7, 8, 9, 10, 12, and 14. After staining, formalin-fixed samples were blindly evaluated for rejection. Reverse transcriptase polymerase chain reaction (rtPCR) using primers specific for beta-actin, IL-2R, IL-6, and TNF was performed on liquid nitrogen-frozen samples. Semiquantitation was accomplished using radionuclide incorporation and beta-scintillation counting. Intestinal histopathology in all isografts (ISO) and POD 3 allografts (ALLO) was normal. Rejection progressed in ALLO from mild on POD 5 to severe by POD 8. rtPCR analysis revealed constitutive expression of IL-2R mRNA in both ISO and ALLO. TNF and IL-6 demonstrated significant increases in mRNA expression in ALLO compared to ISO beginning on POD 5. In summary, intragraft expression of IL-2R mRNA demonstrated late up-regulation in ALLO which did not correlate with rejection. TNF and IL-6 mRNA expression predicted rat SIT rejection. rtPCR analysis of TNF and IL-6 may serve as a useful diagnostic adjunct for rat SIT rejection.

Animals↗
PubMed1994

Adenoviral-mediated gene transfer to bladder in vivo.

This study was designed to examine the potential for gene therapy in bladder in vivo using adenoviral vectors. Gene transfer to rat bladders was accomplished via direct intravesical instillation using a replication-defective adenoviral vector containing a marker gene encoding for Escherichia coli beta-galactosidase (beta-gal). Successful gene transfer was confirmed by analyzing bladder samples for DNA and RNA using polymerase chain reaction (PCR) with primers specific for beta-gal and adeno sequences, detecting beta-gal in full-thickness bladder wall using specific histochemical staining (X-gal) and documenting recombinant protein production. Bladder architecture was preserved, without evidence of distant spread of virus as assessed by PCR. Gene expression was evident for at least 7 days. In summary, bladder cells can be genetically altered using replication-deficient adenoviral vectors via simple intravesical instillation of vector. Introduction of exogenous genetic material is a potentially powerful therapeutic modality for immunomodulation of bladder neoplasms.

Adenoviridae↗
PubMed1994

Current treatment modalities for hepatocellular carcinoma.

OBJECTIVE: This study evaluated the currently available treatment modalities for hepatocellular carcinoma (HCC). SUMMARY BACKGROUND DATA: One of the most common tumors worldwide, HCC has several known risk factors. Untreated HCC typically has a dismal prognosis. Early detection remains the key to successful treatment of this malignancy. Surgical resection has been the mainstay of treatment for HCC, but newer modalities have been recently introduced. METHODS: The authors evaluated the treatment modalities for HCC. RESULTS: Surgical resection affords 5-year survival rates as high as 45% with more favorable subgroups having 1) small tumors, 2) well-differentiated tumors, 3) unifocal tumors, 4) lack of vascular invasion, 5) absence of cirrhosis, and 6) the fibrolamellar variant (FL-HCC). Resection has been limited primarily by low resectability rates and recurrent disease. Newer therapeutic modalities that appear the most promising are transarterial chemoembolization and percutaneous ethanol injection. Neither therapy has been evaluated in a prospective randomized manner. Combination chemotherapy and surgical intervention may provide the best results, but randomized controlled trials with long-term follow-up are needed. As single-treatment modalities, radiation therapy, intravenous chemotherapy, intra-arterial chemotherapy, and immunotherapy play limited palliative roles. CONCLUSIONS: Surgical resection in the form of partial or total hepatectomy is the preferred treatment for HCC. The early detection of tumors by screening high-risk populations is crucial. Randomized trials of combinations of chemotherapy and surgical resection are needed to demonstrate their potential utility for treatment.

Carcinoma, Hepatocellular↗
PubMed1994

One thousand liver transplants. The lessons learned.

OBJECTIVE: To evaluate the first 1000 liver transplants performed at UCLA Medical Center to determine factors responsible for improved results. SUMMARY BACKGROUND DATA: Liver transplant has evolved impressively since the first case was performed in 1963. The 1980s have highlighted this progress with the development of better organ preservation, standardization of operative procedure, improved immunosuppressive agents, and better understanding of patient selection. METHODS: The first 100 consecutive liver transplants (group 1) performed from February 1984 through October 1986 were compared with the last 200 (group 2) performed between September 1991 and June 1992. An analysis was made of donor use; changes in patient candidacy; patient care variables; morbidity and mortality; survival data; and hospital resource use. RESULTS: In group 1, 31% of donors were refused because of medical unsuitability compared with 4% in group 2 (p < 0.0001). In group 1, alcoholic patients comprised 1% of liver transplant candidates compared with 20% group 2 (p < 0.0001). High-risk patients (United Network for Organ Sharing criteria 4) only comprised 11% of patients in group 1 compared with 37% in group 2 (p < 0.0001). Operative time (7.6 hours compared with 5.4 hours), packed cell replacement (17 units compared with 9.5 units), intensive care unit stay (10 days compared with 5 days), and hospital stay (50 days compared with 31 days) were all significantly improved (p < 0.05). In group 1, the 1-year survival rate was 73% and improved to 88% in group 2 (p < 0.0001). CONCLUSIONS: Despite unfavorable donor characteristics (obesity, cause of death, age, hypotension), most organs function well and should not be refused based on history alone. The older and high-risk patient (renal failure, ventilator dependence, portal vein pathology, and so on) is routinely transplanted with good success. Despite liberalization of both donor and recipient criteria, patient survival after liver transplant is improved, use of hospital resources is maximized, and cost reduction is achieved.

Adult↗
PubMed1994

Retroviral-mediated gene transfer into rat experimental liver transplant.

Replication-defective retroviral vectors were used for ex vivo gene transfer into rat liver grafts under conditions mimicking clinical liver transplantation. Supernatant containing single- and double-gene vectors encoding for either the human IL-7 and/or neomycin phosphotransferase genes were used to perfuse the liver grafts during cold ischemia before transplantation. Whole liver grafts were perfused with vector supernatant or medium only. Reduced-size liver grafts (50% hepatectomy) were similarly perfused either immediately after reduction or 24 hr later after induction of active hepatocyte division. After transplantation of these grafts in orthotopic position, the liver tissue was removed at specified intervals, and genomic DNA and mRNA were examined for proviral sequences and expression. Stable integration of the proviral sequences was detected only in reduced-size grafts transplanted 24 hr after hepatectomy. Proviral message of both neomycin phosphotransferase and human IL-7 were present up to 21 days after transduction. This study demonstrates efficient ex vivo gene transfer to donor liver grafts. Gene transfer to livers before transplantation carries the potential to modulate immunogenicity and alter the antigraft immune response.

Animals↗
PubMed1994

Regeneration-induced accelerated rejection in reduced-size liver grafts.

Liver regenerative processes are associated with enhanced expression of alloantigens. Accordingly, we tested the hypothesis that such enhanced surface expression of alloantigens during regeneration of reduced-size liver grafts is associated with accelerated rejection. Our OLT model was LEW (RT1) to BN (RT1n), with donor liver resected by 50%. The study group consisted of reduced-size allografts. Control groups were syngeneic reduced-size isografts and full-size allografts. Reduced-size isograft recipients survived indefinitely. Both isografts and allografts regenerated to their prereduction size within 12 days. Recipients of reduced-size allografts died of accelerated rejection within 12.2 +/- 0.8 days, significantly earlier than recipients receiving full-size allografts (36.2 +/- 4.1 days, P < 0.01). The accelerated rejection in the regenerating allografts was mediated both by cellular and humoral mechanisms, evidenced by earlier lymphocytic invasion of the graft, enhanced donor MHC class II expression, and the emergence of IgM antibodies, directed specifically against donor endothelial antigens. These data suggest that regeneration of reduced-size allografts is accompanied by accelerated cellularly and humorally mediated alloreactivity. Recipients of reduced-size allografts may, therefore, benefit from more potent immunosuppression during the period of active liver regeneration.

Animals↗