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Biomedical subjects

P Ferrara

Publications and source records attributed to P Ferrara.

At least 181 records · Page 10Linked to original sources
PubMed1989

Biochemical analysis of a novel H-2 class I-like glycoprotein expressed in five AKR-(Gross virus) derived spontaneous T cell leukemias.

The H-2 class I Ag profiles of five spontaneous AKR (H-2K) Gross virus leukemic cell lines were analyzed. A novel H-2 class I, "alloantigen"-like glycoprotein was immunoprecipitated and isolated from all the tumor cell lines using an H-2Dd-specific mAb 35-5-8. The novel Ag was also recognized in vitro by anti-H-2Dd-specific CTL. In addition, DNA from all the thymomas, but not the DNA from normal adult AKR thymic cells showed a transcribed gene detectable with an H-2Dd-specific oligonucleotide probe. The molecular profile of the novel antigen was further studied by two-dimensional gel electrophoresis and analyzed by a computer based image analyzer system and reverse-phase HPLC tryptic peptide mapping. Its molecular pattern was different from the syngeneic H-2Kk, H-2Dk, and the allogeneic H-2Dd gene products. The two-dimensional gel pattern of the novel H-2 class I molecule had a different overall structure reflected in isoelectric point, number, and distribution of polypeptide spots. The tryptic peptide map analysis showed six peaks exclusively identified with the novel Ag. The calculated degree of homology with the corresponding H-2Dd, H-Dk, and H-Kk peptides was 41, 56, and 51%, respectively. In addition, an unusual cell surface distribution of the novel Ag was observed in most of the leukemic lines. The removal of sialic acid residues by neuraminidase treatment facilitated the detection of the allodeterminants by anti-H-2Dd-specific mAb and CTL. Furthermore, we showed that in one AKR tumor line, 424, there is a close association of the novel Ag with the syngeneic class I molecules. Prior preclearance of the syngeneic class I molecules revealed the presence of the H-2Dd-like allospecificity. The genetic and molecular relationship between the expression of this novel class I-like glycoprotein and the recently sequenced Q5 gene is under current investigation.

AKR murine leukemia virus↗
PubMed1989

Characterization of a peripheral-type benzodiazepine-binding site in the mitochondria of Chinese hamster ovary cells.

The isoquinoline carboxamide derivative [3H]PK11195, a ligand for the peripheral-type benzodiazepine (BZD) receptor, binds to Chinese hamster ovary (CHO) cell mitochondria in a specific and saturable manner. Scatchard analysis showed the presence of a single-binding site with an apparent dissociation constant (Kd) of 12.0 +/- 1.0 nM and a maximal binding capacity of 23.0 +/- 2.0 pmol/mg protein. The pharmacological characterization of this CHO BZD-binding site, based on the displacement of [3H]PK11195 by several drugs of known binding specificity, indicated that it is of the peripheral-type. The photoaffinity probe [3H]PK14105, a nitrophenyl derivative of [3H]PK11195, specifically labeled a 17 kDa CHO mitochondrial protein. This 17 kDa protein was purified from digitonin-solubilized mitochondria by gel-filtration chromatography and two reverse-phase HPLC steps. The purified material migrated as a single band on silver stained or autoradiographed SDS-polyacrylamide gels, and had an amino acid composition corresponding to a 17 kDa protein rich in Leu, Val, Ala, Gly, and Pro. Analysis of the amino-terminal sequence of the purified 17 kDa protein revealed a blocked amino-terminus.

Affinity Labels↗
PubMed1989

Molecular differences between rat-liver and rat-kidney biliverdin reductase. Implications for their in vivo regulation.

Rat-liver biliverdin reductase exists in two molecular forms. The major form 1 has a molecular mass of 34 kDa, while the minor form 2 has a molecular mass of 56 kDa. Form 1 was converted into a second major form (form 3) with a molecular mass of 68 kDa by a NAD+-dependent peroxisomal dehydrogenase which was induced under conditions of oxidative stress [Frydman, R. B., Tomaro, M. L., Awruch, J. & Frydman, B. (1984) Biochem. Biophys. Res. Commun. 121, 249]. Molecular form 1 from rat kidney was not affected by the dehydrogenase, and a structural explanation for this difference was therefore sought. Both form 1 biliverdin reductases, isolated from rat liver and kidney, were purified to homogeneity using affinity chromatography, FPLC and HPLC techniques. The homogeneous enzymes were found to be identical when compared by their HPLC retention times, amino acid compositions and electrophoretic behaviour on polyacrylamide gels under non-denaturing conditions and on SDS/polyacrylamide gels. On HPLC analysis the peptides resulting from the CNBr cleavage were found to be the same for both enzymes, when either the native enzymes or their thioethylpyridine derivatives were compared. When the HPLC fingerprints of the tryptic digests were compared, they were found to be very similar, except for a peptide eluting at 31.60 min in the liver digest and at 23.60 min in the kidney digest. When the enzyme from both origins was alkylated with 4-dimethylaminoazobenzene-4'-iodoacetamide and then digested with trypsin, the HPLC fingerprints of the alkylated cysteine-carrying peptides were almost identical, except for a peptide with a retention time of 19.03 min in the liver digest and of 18.19 min in the kidney digest. The liver reductase was not amenable to Edman degradation suggesting a block at the NH2-terminus; in the kidney enzyme, however, it was free and an NH2-terminal sequence of 12 amino acids could be determined. The liver enzyme was found to be more sensitive toward p-hydroxymercuriphenyl sulfonate than the kidney enzyme.

Alkylation↗
PubMed1989

The impact of radioimmunoguided surgery (RIGS) on surgical decision-making in colorectal cancer.

Radioimmunoguided surgery (RIGS system) was performed in ten patients with rectal or low sigmoid colon carcinoma with the use of a hand-held gamma detector (Neoprobe 1000) intraoperatively and externally after injection of radiolabeled (125I) monoclonal antibody to detect pelvic and metastatic tumor. Fifteen procedures, including six exploratory laparotomies, four transperineal explorations, two transsacral explorations, one transvaginal biopsy, one brachytherapy, and one transanal polypectomy, were performed. Two patients had previous low anterior resection, seven abdominoperineal resection, and one a rectal polypectomy. Five patients had previous pelvic radiation therapy. Reoperation was indicated by elevated CEA levels in seven patients (70 percent), persistent pelvic pain in six (60 percent), and a suspicious radiologic study in seven (70 percent). RIGS system localized tumors verified by histopatholoy in all ten patients (100 percent); one patient with a positive CT scan and probe findings lacked histopathologic confirmation on frozen section, but had a tumor confirmed on permanent histology. Five major abdominal operations were avoided; in five patients major modifications were made in the surgical procedure based on probe findings. Six received chemotherapy or radiation therapy based on findings of the RIGS system. In six patients with negative or equivocal CT scans, the RIGS system localized histopathologically confirmed tumor. Major abdominal procedures can be avoided, the surgical approach modified, and other modes of therapy instituted earlier with the use of the RIGS system.

Aged↗
PubMed1989

A rapid two-step purification of rat cystatin C, one major inhibitor of cysteine proteinases.

Rat cystatin C was purified to apparent homogeneity from rat urine after induction of a tubular dysfunction with sodium chromate. Twentyfold concentrated urine was chromatographed by a rapid purification procedure. A two-step purification including affinity chromatography on carboxymethyl papain- Sepharose and high-resolution anion exchange chromatography was developed. The purified protein has an apparent molecular mass of 15 kDa and pI of 10.2; its aminoacid composition was similar to human cystatin C. As opposed to previous data, purified urinary rat cystatin C did not contain significant amounts of carbohydrate. Antisera against rat cystatin C, raised in rabbits, partially cross-reacted with human and mouse cystatin C, indicating their antigenic similarities. Like human cystatin C, native rat cystatin C, named slow form, is degraded into a more acidic form, called fast form, by a loss of N-terminal amino acids; fast form displayed a pI of 9.4.

Amino Acid Sequence↗
PubMed1989

[High-risk focal breast lesions in echography and mammography].

Proliferative breast diseases include a group of lesions which occupy an intermediate position between benign and malignant lesions and are extremely likely to develop into carcinomas. The authors studied 81 patients who had been surgically biopsied on the basis of mammographic and/or US findings. In 33/81 (40.7%) of them proliferative lesions were diagnosed at histology. Mammography was able to identify 18/33 lesions (54.5%) versus US 26/33 (81.8%). On the basis of these findings, a group of patients at risk for cancer could be identified. As a matter of fact, proliferative lesions, especially those presenting with atypical hyperplasia, are associated with a higher risk of developing into carcinomas than those presenting with typical features. For these patients, the authors suggest more frequent clinical and instrumental screening.

Adult↗
PubMed1989

Radioimmunoguided surgery in recurrent colorectal cancer: the role of carcinoembryonic antigen, computerized tomography, and physical examination.

From January 1986 to December 1987, 32 patients with recurrent colorectal cancer had second-look radioimmunoguided surgery (RIGS system). All patients had pathologic confirmation of recurrence. The RIGS system identified 81% of recurrences, and in six patients recurrent tumor was identified only by RIGS. All patients had physical examination, carcinoembryonic antigen (CEA) assay, and computerized tomography of the abdomen and pelvis. Detection of recurrence was based on symptoms in six, elevated CEA value in 25, and physical examination in one. The CEA was elevated preoperatively in 30 patients; two false-negative results occurred in symptomatic patients who had pelvic recurrence. The median CEA value in those with liver recurrence was 30 ng/ml (range 5.2 to 298) and for pelvic recurrence 13 ng/ml (range 1.9 to 31) (P less than .05). The overall sensitivity of CT was 41% (abdomen other than liver 37%, liver 56%, and pelvis 22%). The combination of elevated CEA, symptoms, and physical findings identified 100% of recurrences. We conclude that a rising CEA remains the most accurate indicator of recurrence. CT should not be done routinely to detect recurrent colorectal cancer unless CEA is elevated or the patient is symptomatic. In our study the intraoperative use of the RIGS system aided the surgeon in identifying occult tumors.

Adolescent↗
PubMed1988

Trichokirin, a ribosome-inactivating protein from the seeds of Trichosanthes kirilowii Maximowicz. Purification, partial characterization and use for preparation of immunotoxins.

A protein, here named trichokirin, was extracted from the seeds of Trichosanthes kirilowii and purified by ion-exchange and gel-filtration chromatography. Trichokirin is a basic glycoprotein of apparent relative molecular mass of 27,000 with a strong ribosome-inactivating activity. Alignment of the trichokirin, trichosanthin and momordin N-terminal sequences shows a substantial degree of homology. Trichokirin was conjugated to a monoclonal antibody directed against the Thy 1.2 antigen with the cleavable dimethyl 3,3'-dithiobispropionimidate cross-linking reagent. This immunotoxin selectively killed leukemia cells expressing the Thy 1.2 antigen. The addition of ammonium chloride, which increases the cytotoxicity of ricin A-chain immunotoxins, blocks that of the trichokirin immunotoxin, suggesting that they enter cells by different mechanisms. In vivo studies showed that the pharmacokinetic properties of the trichokirin immunotoxin could be more advantageous than those of the ricin A-chain immunotoxins for in vivo applications.

Amino Acid Sequence↗
PubMed1988

Role of the N-terminus of rat pheochromocytoma tyrosine hydroxylase in the regulation of the enzyme's activity.

Activation of rat pheochromocytoma tyrosine hydroxylase by limited tryptic proteolysis was investigated. The modifications produced upon the enzyme's structure were analyzed with the use of sodium dodecyl sulfate/polyacrylamide gel electrophoresis and tyrosine hydroxylase activity was measured all through the digestion. During the proteolysis the activity of tyrosine hydroxylase was elevated threefold at the same time as a 56-kDa tryptic fragment was formed. When the enzyme was phosphorylated, at its N-terminal region, by a kinase copurified with tyrosine hydroxylase, the major 56-kDa species did not appear to be phosphorylated on the autoradiograph, suggesting that it was derived from the native subunit by cleavage of the N-terminal of the protein. The reactivity of the 2/40/15 anti-(tyrosine hydroxylase) monoclonal antibody with the N-terminal of tyrosine hydroxylase was also investigated, using the Western-blot technique. This antibody reacted with the 62-kDa hydroxylase subunit but not with the 60-kDa tryptic fragment; the amino acid sequences of these two species showed that the 60-kDa fragment lacked the first 16 N-terminal amino acids of the native molecule. These results suggest that the N-terminal region of tyrosine hydroxylase is apparently responsible for an inhibition of the hydroxylase activity and that the first N-terminal amino acids of the hydroxylase are necessary for the recognition of the enzyme by its antibody.

Amino Acid Sequence↗
PubMed1988

Biological activity of chitosan: ultrastructural study.

Reparative processes are reconstructive phenomena in which cellular elements and extracellular matrix glycoproteins interact to build the injured tissue. Biomaterials can be used to improve or stimulate reconstruction. In the present experimental investigation, tissue repair induced by chitosan, an 86.8% deacetylated poly(GlcNAc), was monitored by morphological analysis. To evaluate its biological role, chitosan was positioned in contact with dura mater or was used as a dura mater substitute. This polysaccharide, having structural characteristics similar to glycosaminoglycans, seems to mimic their functional behaviour. The inductive and stimulatory activity of chitosan on connective tissue-rebuilding is clearly demonstrated, and it is suggested that chitosan could be considered a primer on which a normal tissue architecture is organized.

Animals↗
PubMed1988

Cloning and sequencing of the 52K cathepsin D complementary deoxyribonucleic acid of MCF7 breast cancer cells and mapping on chromosome 11.

Two lambda gt11 libraries containing complementary DNAs from human breast cancer MCF7 cells were screened by expression with monoclonal antibodies to the secreted 52K protein and with a 36-mer oligonucleotide derived from the N-terminal amino acid sequence of the secreted 52K protein. Four overlapping clones were sequenced, and found to be extensively homologous to the cathepsin D of normal human kidney, except for 5-point mutations resulting in one amino acid change (Ala to Val) in the profragment of cathepsin D. Northern blot analysis showed the 2.2 kilobase (kb) cathepsin D mRNA to be induced by estradiol in MCF7 cells and produced constitutively at high levels in the estrogen-receptor-negative BT20 cell line. A simple restriction pattern consistent with the restriction map of cathepsin D cDNA was obtained in Southern blot analysis of MCF7 cell DNA. In situ hybridization of the 52K-9 cDNA probe on normal lymphocytes assigned the 52K cathepsin D gene at the extremity of the short arm of chromosome 11, in the p15 band, close to the H-ras gene and in the region whose deletion increases the risk of invasive breast cancer. We conclude that the estrogen induced 52K protein has the same sequence as normal pro-cathepsin D and we propose that the 52K protein correspond to the only pro-cathepsin D expressed in MCF7 cells.

Breast Neoplasms↗
PubMed1988

Effect of general anesthesia on syncytiotrophoblast plasma membranes from human placenta.

This investigation shows that a general anesthetic produces similar effects in vivo and in vitro. Anesthesia with a barbituric drug, Thiopental, induces an increase in membrane fluidity and a decrease in the activity of acetylcholinesterase in syncytiotrophoblast plasma membranes (SPM) obtained from placentas after caesarean section. The same effects can be reproduced in vitro after anesthetic addition to the isolated plasma membranes. Morphological and freeze-fracturing studies also suggest that membrane protein components are affected by anesthetics.

Acetylcholinesterase↗
PubMed1988

[Audio-vestibular changes in patients with rheumatoid arthritis].

A functional audio-vestibular investigation based on impedance metric techniques and electronystagmography was carried out in a group of patients with "classical" rheumatoid arthritis (RA) and with treated or untreated "definite" RA in various stages. Data obtained from these patients were compared with those obtained from a control group. Significant hypoacusis of the transmissive type was found in initial stages of RA while sensorineural or mixed type hypoacusis was found in later stages of RA. Significant vestibular alterations of the central type suggesting supratentorial involvement were found in several cases independently of the stage of RA and of the age of the patients.

Acoustic Impedance Tests↗