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R Shapira

Publications and source records attributed to R Shapira.

At least 19 recordsLinked to original sources
PubMed2001

Tyrosine cross-linking of extracellular matrix is catalyzed by Duox, a multidomain oxidase/peroxidase with homology to the phagocyte oxidase subunit gp91phox.

High molecular weight homologues of gp91phox, the superoxide-generating subunit of phagocyte nicotinamide adenine dinucleotide phosphate (NADPH)-oxidase, have been identified in human (h) and Caenorhabditis elegans (Ce), and are termed Duox for "dual oxidase" because they have both a peroxidase homology domain and a gp91phox domain. A topology model predicts that the enzyme will utilize cytosolic NADPH to generate reactive oxygen, but the function of the ecto peroxidase domain was unknown. Ce-Duox1 is expressed in hypodermal cells underlying the cuticle of larval animals. To investigate function, RNA interference (RNAi) was carried out in C. elegans. RNAi animals showed complex phenotypes similar to those described previously in mutations in collagen biosynthesis that are known to affect the cuticle, an extracellular matrix. Electron micrographs showed gross abnormalities in the cuticle of RNAi animals. In cuticle, collagen and other proteins are cross-linked via di- and trityrosine linkages, and these linkages were absent in RNAi animals. The expressed peroxidase domains of both Ce-Duox1 and h-Duox showed peroxidase activity and catalyzed cross-linking of free tyrosine ethyl ester. Thus, Ce-Duox catalyzes the cross-linking of tyrosine residues involved in the stabilization of cuticular extracellular matrix.

Amino Acid Sequence↗
PubMed2001

Efficacy of lamivudine for the treatment of hepatitis B virus infection after liver transplantation in children.

BACKGROUND: There is at present very little information about hepatitis B virus (HBV) infection in children after liver transplantation. This is the first study to assess the safety and efficacy of lamivudine in this patient population. METHODS: We describe three children aged 5-14 years who underwent liver transplantation for fulminant hepatitis A, hyperoxaluria, and cystic fibrosis. Despite adequate immunoprophylaxis, two of the children who were serum hepatitis B surface antigen-positive before transplantation (HBV DNA-negative by hybridization) had a reactivation of the disease, and one had a de novo HBV infection, at 12-18 months after transplantation. Lamivudine 3 mg/kg was administered on a compassionate-use basis for 14-36 months. RESULTS: After 1 month of therapy, HBV DNA disappeared from the serum in all patients by hybridization and in two patients by polymerase chain reaction. In all three children, alanine transaminase levels normalized. One child developed lamivudine resistance after 22 months with no evidence of hepatic decompensation. Repeated liver histological studies revealed progression of hepatic fibrosis in one child. All children remained serum hepatitis B surface antigen- and hepatitis B e antigen-positive. No adverse effects of the drug were noted. CONCLUSION: Lamivudine is beneficial and well tolerated in children with HBV infection after liver transplantation.

2-Aminopurine↗
PubMed2001

[Liver failure with coagulopathy in an infant with tyrosinemia].

Tyrosinemia is an inherited autosomal recessive condition. We present a 5 week-old boy with this disorder. He was admitted because of a fever, vomiting and lethargy. The laboratory tests confirmed a coagulopathy with prolonged prothrombin time (PT), partial thromboplastin time (PTT) and a decreased serum fibrinogen. The alpha-fetoprotein level was markedly elevated. To confirm the diagnosis of tyrosinemia, quantitative urinary succinylacetone was measured. Although overt liver failure with coagulopathy may be part of the representation of tyrosinemia, a significant coagulopathy in the absence of overt signs of liver disease has not been emphasized as a clue to the diagnosis of this condition.

Biomarkers↗
PubMed2000

Cloning and expression of the genes involved in the production of and immunity against the bacteriocin lacticin RM.

The production of lacticin RM, a novel bacteriocin produced by Lactococcus lactis subsp. lactis EZ26, is associated with the presence of a 6-kb plasmid, pHU1. The information necessary for lacticin RM production and immunity was localized to a 2.5-kb SalI-Eco47III fragment. Sequencing analysis of this fragment revealed the presence of six open reading frames (ORFs). Deletion and mutation analyses showed that orfX and orfY are not required for lacticin RM production or immunity, whereas the other ORFs (lacA, lacF, lacG and lacI) are necessary for the bacteriocin's production. Transcription analysis indicated that lacA, lacF and lacG are organized in an operon. lacA is probably the lacticin RM structural gene. It putatively encodes a 134-amino acid peptide, and it does not share homology with known bacteriocins. The deduced LacG protein is hydrophobic and consists of six potential trans-membrane helices. lacF encodes a conserved ATP-binding domain homologous to ABC transporters known in bacteriocin immunity systems. LacF and LacG may form an active ABC transporter. Gene-disruption mutations indicated that both are required for immunity against lacticin RM. lacI encodes a small cationic protein, which is required for the production of and immunity to lacticin RM. Protection was obtained only when lacF, lacG and lacI were present together.

ATP-Binding Cassette Transporters↗
PubMed2000

[Experience with 100 liver transplant recipients at the Rabin Medical Center and the Schneider Children's Medical Center].

Liver transplantation is the treatment of choice for end-stage liver disease. During the past 8 years we performed 102 liver transplants in 84 adults and 16 children. In the adults, 9 were combined transplants: 1 a liver-pancreas transplant for type I diabetes, and 8 liver-kidney transplants. In the children, transplants included 5 whole-livers, 5 left-lateral liver segments from living-related donors, 4 reduced-grafts of right or left lobes, and 2 split left-lateral segments. At a mean follow-up of 31 months (range 1-96) 70 were alive, 3 had died during surgery and 15 during the first postoperative months. Mortality was due to primary graft non-function (7), sepsis (10), intracranial hemorrhage (1), tumors (4), recurrent hepatitis B (2), biliary strictures (2) and chronic rejection (1). The 1- and 4-year survival rates were 79.5% and 69.6%, respectively. After transplantation, 10 developed biliary stricture (5 corrected by balloon dilatation) and 8 anastomotic stricture (7 corrected by surgery), and there were 2 multiple intrahepatic strictures. There was hepatic artery thrombosis in 5, including 4 children. In 3, grafts were salvaged by thrombectomy and 2 others underwent re-transplantation. In those who survived transplantation by more than 1-month, recurrent hepatitis B was seen in 6 of 17 (35%) and recurrent hepatitis C in 12 of 19 (63%). Thus, results of our first 100 liver transplants are similar to those reported by larger centers, showing that in an appropriate setting good results can be achieved by small transplant programs.

Adolescent↗
PubMed1999

Hepatic artery thrombosis in pediatric liver transplantation: graft salvage after thrombectomy.

Hepatic artery thrombosis (HAT) is a devastating complication that may occur after orthotopic liver transplantation (OLT). A higher incidence has been reported in children. Salvage of the graft by thrombectomy has been suggested as an alternative to re-transplantation. In this study we report the outcome of three children who underwent thrombectomy for HAT. Between January 1992 and June 1998, 14 children (< 17 yrs of age) underwent liver transplantation. Three developed HAT (one a whole-liver graft recipient, age 17; two living-related graft recipients, ages 4 and 4.5 yr). In the first patient, thrombosis of the hepatic artery was associated with scattered areas of parenchymal necrosis on computed tomography. In the two living-related patients, HAT was found incidentally during re-exploration for bleeding (day 2 and day 10). Thrombectomy was performed in all three patients. At 18-24 months after thrombectomy, all three children had normal graft function. In the first patient, complete regeneration of the liver has been documented by computed tomography and a late asymptomatic recurrent thrombosis is suggested by absence of arterial flow on Doppler examination. The hepatic artery is patent in the two living-related recipients. One of these living-related recipients developed ischemic bile duct stricture and underwent successful percutaneous balloon dilatation. We conclude that long-term normal graft function can be achieved by thrombectomy in pediatric liver recipients with HAT, even in the presence of limited parenchymal damage.

Adolescent↗
PubMed1999

Retrospective review of cystic fibrosis presenting as infantile liver disease.

The mode of presentation, clinical course, and outcome of 12 infants with cystic fibrosis and liver disease referred over an 18 year period were investigated retrospectively. Median age at presentation was 6.5 weeks (range, 5-12). Two thirds were boys. Conjugated hyperbilirubinaemia was the presenting symptom in 11 patients, and hypoalbuminaemia in one. Jaundice was cleared over a median period of 7.36 months. Eight patients had bile duct proliferation on liver biopsy and one required cholangiography to exclude biliary atresia. Classic histological features of cystic fibrosis were only present in two children biopsied at 8 and 18 months. Three patients had meconium ileus, including one infant with concomitant alpha(1) antitrypsin deficiency, who required early liver transplantation. All other patients had no signs of significant chronic liver disease during a median follow up of 42 months (range, 10-205). Children with cystic fibrosis and infantile liver disease have a good short and medium term prognosis.

Albuminuria↗
PubMed1999

Possible synergistic effect of nisin and propionic acid on the growth of the mycotoxigenic fungi Aspergillus parasiticus, Aspergillus ochraceus, and Fusarium moniliforme.

The effects of nisin and propionic acid (PA) on aflatoxin production and on mycelial growth and spore germination of the mycotoxigenic fungi Aspergillus parasiticus, A. ochraceus, and Fusarium moniliforme were investigated. The growth of A. ochraceus was completely inhibited on media containing PA with nisin in concentrations of 0.05% PA with 1,000 ppm nisin, and 0.1% PA with 500 or 1,000 ppm nisin. The growth of both F. moniliforme and A. parasiticus was completely inhibited by PA with nisin at a concentration of 0.1% PA with 1,000 ppm nisin. Nisin alone caused a significant increase in mycelial growth when applied to A. ochraceus at 500 or 1,000 ppm and when applied to A. parasiticus at 1,000 ppm. Spore germination of A. ochraceus was completely inhibited on media containing 0.1% PA with 500 or 1,000 ppm nisin. Spores of F. moniliforme failed to germinate in 0.05% PA with 500 or 1,000 ppm nisin, whereas spores of A. parasiticus did not germinate on media containing 0.1% PA with 1,000 ppm nisin. For all three fungi tested, the inhibitory effect on mycelial growth was found to be fungistatic rather than fungicidal. The combined treatment of PA with nisin produced better fungistatic activity than treatment involving either material alone. Nisin, applied alone, did not stimulate aflatoxin production (expressed by microg toxin/mg mycelium), but the combined treatment at certain concentrations was inhibitory to aflatoxin B1 or G1. The production of aflatoxin G1, but not of B1, was stimulated in 0.05% PA with 1,000 ppm nisin and on media containing 0.1% PA with 100 ppm nisin. Nisin is currently applied in foods to prevent spoilage induced by bacteria but not by mold. The results of the present study indicate that a combined treatment of nisin in small concentrations of PA might be useful in preventing mold damage in certain foods and stored grain.

Aflatoxins↗
PubMed1998

[Living-related liver transplantation--first experiences at Rabin Medical Center].

Our experience with living-related liver transplantation is described. In 2 boys and 1 girl, aged 4-4.5 years with acute, fulminating hepatitis A, the presence of very severe jaundice (bilirubin levels > 18 mg%) associated with severe coagulopathy (INR > 10) and encephalopathy indicated the need for urgent liver transplantation. In all 3 cases the left lateral hepatic segment of a matched blood type parent was transplanted. None of the donors suffered a serious complication postoperatively and all returned to full activity in 6-16 weeks. The post-transplantation course was uneventful in 1 child, but in the other 2 there was hepatic arterial thrombosis in 1 at 1 day and in the other at 8 days post-transplantation. Early detection of arterial thrombosis by Doppler sonography permitted salvage of the 2 hepatic grafts after thrombectomy and re-anastomosis. In 1 of these 2 children an anastomotic biliary stricture was found 2 months after transplantation. It was corrected at surgery and a percutaneous stent was inserted. All 3 children are alive with normal graft function at 2, 7 and 8 months post-transplantation, respectively. This initial experience indicates that living-related liver transplantation is feasible in Israel. The technique might help to solve our severe organ shortage for children awaiting liver transplantation.

Adult↗
PubMed1997

Molecular characterization of the replicon of the Pediococcus pentosaceus 43200 pediocin A plasmid pMD136.

The pediocin A-encoding plasmid of Pediococcus pentosaceus 43200, pMD136, was characterized by restriction enzyme analysis. Analysis of its replicon was facilitated by the construction of a probe vector consisting of the Escherichia coli plasmid pSP72 and the cat gene from Staphylococcus aureus plasmid pC194. The replication region of pMD136 was localized on a 1.6-kb EcoRI/BglII fragment. Sequencing analysis revealed a non-coding region, repA, spanning the first 440 bp, followed by an open reading frame, repB, encoding a putative protein of 390 amino acids. The non-coding region contained two sets of 6-bp and two sets of 22-bp direct repeats and two sets of inverted repeats upstream of the open reading frame. Strong homology of the isolated replicon was found to theta-type replicons of Lactococcus lactis plasmids. Segregational stability assay suggested at least two regions as potentially involved in the stabilization of pMD136. The plasmid's strong homology to other theta-type replicons and its relatively high stability suggest that pMD136 belongs to the widespread family of theta-replication plasmids.

Amino Acid Sequence↗
PubMed1997

Functional characterization of pediocin PA-1 binding to liposomes in the absence of a protein receptor and its relationship to a predicted tertiary structure.

The physicochemical interaction of pediocin PA-1 with target membranes was characterized using lipid vesicles made from the total lipids extracted from Listeria monocytogenes. Pediocin PA-1 caused the time- and concentration-dependent release of entrapped carboxyfluorescein (CF) from the vesicles. The pediocin-induced CF efflux rates were higher under acidic conditions than under neutral and alkaline conditions and were dependent on both pediocin and lipid concentrations. A binding isotherm constructed on the basis of the Langmuir isotherm gave an apparent binding constant of 1.4 x 10(7) M-1 at pH 6.0. The imposition of a transmembrane potential (inside negative) increased the CF efflux rate by 88%. Pediocin PA-1 also permeablized synthetic vesicles composed only of phosphatidylcholine. Sequence alignments and secondary-structure predictions for the N terminus of pediocin PA-1 and other class IIa bacteriocins predicted that pediocin PA-1 contained two beta-sheets maintained in a hairpin conformation stabilized by a disulfide bridge. The structural model also revealed patches of positively charged residues, consistent with the argument that electrostatic interactions play an important role in the binding of pediocin PA-1 to the lipid vesicles. This study demonstrates that pediocin PA-1 can function in the absence of a protein receptor and provides a structural model consistent with these results.

Amino Acid Sequence↗
PubMed1997

Development of polyclonal antibodies for detection of aflatoxigenic molds involving culture filtrate and chimeric proteins expressed in Escherichia coli.

Polyclonal antibodies (PAb) were raised against an aflatoxigenic strain of Aspergillus parasiticus by using two different sources for antibody elicitation: (i) filtrate of a culture on which the fungus had been grown (ii) and two chimeric proteins, expressed in Escherichia coli as separate products, of the genes ver-1 and apa-2, which are involved in aflatoxin biosynthesis. The gene products were amplified by PCR, and each was cloned into the E. coli expression vector pGEX2T. Upon induction, the bacteria overexpressed 38- and 33-kDa chimeric proteins corresponding to the N-terminal domains of the genes ver-1 and apa-2, respectively. The chimeric proteins were isolated and affinity purified for use as antigens. The specificity of the raised antibodies was examined by enzyme-linked immunosorbent assay (ELISA). The PAbs raised against the culture filtrate reacted with all the species of Aspergillus and Penicillium tested but not with Fusarium species or corn gain. However, the PAbs elicited against the chimeric proteins were highly specific, showing significantly higher ELISA absorbance values (A405) against A. parasiticus and A. flavus than against the other fungi tested and the corn grain. The approach of utilizing gene products associated with aflatoxin biosynthesis for antibody production therefore appears to be feasible. Such a multiantibody system combined with the PCR technique, could provide a useful tool for the rapid, sensitive, and accurate detection of aflatoxin producers present in grains and foods.

Aflatoxin B1↗
PubMed1997

Large-scale preparation and characterization of recombinant ovine placental lactogen.

To clone ovine placental lactogen (oPL) cDNA, total RNA from sheep placental cotyledon was reverse transcribed and the single-stranded cDNA was PCR-amplified with 5' and 3' primers containing, respectively, NcoI and PstI sites. The oPL cDNA fragment amplified between these two primers extended from A(-1) to the natural stop codon. The PCR product was gel-purified and subcloned into a Puc vector and the insert was sequenced on both strands, revealing several differences relative to the published sequence: S19N, S69N, D129E and R165Q. We assume that these differences can be accounted for by the high level of individual polymorphism, which has been described in detail for PLs of different species. The insert was subcloned into NcoI/ PstI-digested pTrc99A procaryotic expression plasmid and protein expression was induced by isopropyl-1-thio-beta-D-galactopyranoside. Because of low expression, oPL's cDNA was further subcloned into pET8 procaryotic expression plasmid. Its expression in E. coli strain BL21 transformed with this vector yielded 30-40 mg/l. The expressed protein, found in the inclusion bodies, was refolded into a monomer and purified on a Q-Sepharose column to homogeneity. Structural analysis using circular dichroism revealed a spectrum similar to that of human GH (hGH) thereby indicating proper refolding. Gel filtration and binding experiments, including real-time kinetic measurements using the surface plasmon resonance method revealed that oPL forms transient homodimeric complexes with extracellular domains of prolactin receptors from rabbit, rat and bovine and with hGH receptor. The purified oPL was biologically active in an Nb2-11C cell proliferation bioassay, in its ability to stimulate beta-casein synthesis in explants of ovine and rabbit mammary gland and fat synthesis in explants of bovine mammary gland, and in a proliferation assay using FDC-P1 cells transfected with rabbit or hGH receptors.

Animals↗