Biomedical subjects

R Baumal

Publications and source records attributed to R Baumal.

At least 37 records · Page 2Linked to original sources

PubMed1990

Cloning and expression of the human S100 beta gene.

S100 protein is a low molecular weight, EF-hand, Ca2(+)-binding protein widely distributed and conserved in the central nervous system of vertebrates. The gene coding for the beta subunit of human S100 protein (S100 beta) has been recently mapped to chromosome 21. In order to study the expression of this gene in normal and abnormal brain development, we have isolated and characterized overlapping genomic clones spanning the region coding for human S100 beta and its flanking sequences. The intron-exon organization of the human S100 beta gene is similar to that of the genes coding for several other members of the S100 protein subfamily of EF-hand proteins. The human S100 beta gene is composed of 3 exons, the first of which specifies the 5'-untranslated region, while the second and third each encode a single EF-hand, Ca2(+)-binding domain. The promoter region contains several potential regulatory transcription elements including the cAMP-responsive elements CRE and AP-2. A novel sequence motif, the S100 protein element, situated in close proximity to the TATA box of the genes of several members of the S100 protein subfamily, has been identified. In addition, multiple repeats with similar nucleotide sequence and location to the recently reported beta globin direct repeat elements have been also found in the human S100 beta promoter. A full length (17.3 kilobases) copy of the human S100 beta gene was constructed and transfected into rat glioma C6 cells. Stable transfectants were shown to express correctly initiated transcripts of the human S100 beta gene, indicating that the cloned sequences contain functional regulatory transcription elements.

Amino Acid Sequence↗

PubMed1990

Inhibition of human tumor growth by intraperitoneal immunotoxins in nude mice.

Intracavitary administration of immunotoxins may play a role in the control of malignant effusions. Selection of immunotoxins for this form of therapy is based on their prior evaluation in preclinical studies. Monoclonal antibodies (mAb) 454A12 (antitransferrin receptor), and 260F9 are directed against antigens which are present on tumor cells in pleural and peritoneal effusions of patients with adenocarcinoma of the breast and ovary. In the present study, immunotoxins derived by conjugating these mAb to recombinant ricin A (rRA) were shown to be cytotoxic to human ovarian adenocarcinoma HEY cells in vitro and in vivo. In the in vitro assay 454A12-rRA and 260F9-rRA were 1000-fold and 10-fold, respectively, more cytotoxic than free rRa against HEY cells, and both immunotoxins were potentiated approximately 1000-fold by monensin. For in vivo studies HEY cells were injected i.p. into nude mice at a challenge dose (3 x 10(5) cells) which produced carcinomatosis with ascites, leading to death 30 days following injection. Administration of 454A12-rRA i.p. following the challenge dose resulted in a complete cure, whereas administration of 260 F9-rRA with monensin significantly prolonged survival. The greater cytotoxicity of 454A12-rRA than 260F9-rRA against HEY cells could be accounted for by the greater number of binding sites and higher internalization rate for 454A12-rRA and mAb 454A12 than 260F9-rRA and mAb 260F9, respectively. These results suggest a potential role for 454A12-rRA and 260F9-rRA plus monensin in the intracavitary therapy of malignant effusions associated with carcinoma of breast and ovary. In the case of 260F9-rRA, this represents the first preliminary indication of the suitability of this immunotoxin for intracavitary therapy of malignancies.

Adenocarcinoma↗

PubMed1990

A study by immunofluorescence microscopy of the NC1 domain of collagen type IV in glomerular basement membranes of two patients with hereditary nephritis.

The NC1 domain of the collagen type IV molecule, the major component of glomerular basement membranes (GBM), consists of dimers and 24 kilodalton (K), 26 K and 28 K monomers in man, and contains the Goodpasture antigen. Serum obtained from patients with Goodpasture's syndrome has been reported not to stain GBM of most male and some female patients with hereditary nephritis (HN) by immunofluorescence (IF) microscopy. In the present study, GBM seen on the renal biopsies of 2 patients (one male and one female) with HN were examined by IF to ascertain whether NC1 monomers were detectable. Three reagents were used: a plasmapheresis fluid (PPF) obtained from a patient who was treated for anti-GBM nephritis (human anti-GBM PPF); a commercial rabbit antibody against human NC1; and a rabbit antibody raised by us against dog NC1, which cross-reacted with human NC1. All 3 reagents detected NC1 determinants in GBM of normal human kidney by IF and reacted with human NC1 by a plate-binding radioimmunoassay (RIA). The human anti-GBM PPF bound to 28 K and 26 K monomer components of NC1 by Western blotting, the rabbit anti-human NC1 antibody bound to 26 K and 24 K monomers, while the rabbit anti-dog NC1 antibody bound only to the 26 K monomer. By IF, the human anti-GBM PPF did not stain GBM of the male patient with HN, but produced segmental staining of GBM (i.e., some GBM stained, while others did not) of the female patient. In contrast, the rabbit anti-NC1 antibodies produced global staining by IF of GBM of both patients. The absence of staining (i.e., global or segmental) seen with the human anti-GBM PPF implied that the 26 K and 28 K monomers of NC1 were either absent from GBM, or were present but altered structurally, leading to a diminution in their immunological reactivity. However, the positive staining observed with the rabbit anti-NC1 antibodies implied that the 26 K monomer was actually present in GBM. Hence, we postulate that the 26 K monomer of NC1 in GBM was structurally altered, and that the 28 K monomer was either absent, or present but altered. These findings suggest that there is an abnormality of more than one monomer of NC1 in GBM of patients with HN.

Basement Membrane↗

PubMed1990

S100 protein expression in human melanoma cells: comparison of levels of expression among different cell lines and individual cells in different phases of the cell cycle.

The synthesis of S100 protein in cultured human melanoma cells was examined using metabolic labeling with [35S]methionine, immunoprecipitation with anti-S100 protein antiserum, and polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. Six of seven cell lines derived from melanomas synthesized relatively large amounts of S100 protein, whereas three cell lines derived from normal melanocytes synthesized lesser amounts. Synthesis of S100 protein was not detected in 10 human cell lines of nonneuroectodermal origin. Analysis of poly(A+) RNA from one melanoma cell line by Northern blot hybridization with a probe specific for the beta subunit of rat S100 protein revealed a single mRNA species of 1.0 kb coding for the human protein. Flow cytometric analysis of individual cells of two melanoma cell lines and the rat glioma cell line C6 indicated that G0/G1 cells were heterogeneous with respect to S100 protein expression, while almost all the cells in S + G2 + M expressed S100 protein. These results suggest that expression of S100 protein in G0/G1 could be a prerequisite for progression of the cells through the cell cycle.

Blotting, Northern↗

PubMed1990

Renal calcium handling in cystic fibrosis: lack of evidence for a primary renal defect.

Because cystic fibrosis (CF) epithelia have ion transport abnormalities that may in part be regulated by intracellular calcium metabolism, and the kidney is actively involved in both ion transport and calcium homeostasis, we have investigated renal calcium handling in CF. Twenty-four-hour urine collections were analyzed in 34 CF patients (age 5 to 35 years) and kidney ultrasound studies were performed in 17 CF patients (age 6 months to 23 years). Renal histologic findings at postmortem examination of 14 CF patients (age 4 months to 23 years) were compared with those of 12 patients (age 11 months to 17 years) with other chronic illnesses (6 congenital heart disease, 6 malignancy). In 30 of the 34 CF patients urinary calcium excretion was normal (less than 4 mg (0.1 mmol)/kg/24 hr). Four CF patients had hypercalciuria (calcium excretion 4.4 to 8.8 mg (0.11 to 0.22 mmol)/kg/24 hr). However, these patients had other possible explanations for hypercalciuria, such as immobilization (n = 2), increased dietary sodium load (n = 1), and glucocorticoid therapy (n = 1). None of the 17 patients examined by renal ultrasonography had nephrocalcinosis. Five CF patients had histologic evidence of sparse nephrocalcinosis at autopsy. However, 6 of 12 autopsy kidney specimens from patients with other chronic illnesses and similar preterminal events also showed nephrocalcinosis. The hypercalciuria and nephrocalcinosis in CF and other chronic debilitating diseases may be explained by factors known to affect calcium handling. Our evidence does not support a primary renal defect as the basis of hypercalciuria and nephrocalcinosis in CF.

Adolescent↗

PubMed1989

Acute changes in renal function associated with deferoxamine therapy.

In three patients who received intravenous deferoxamine there was a twofold to eightfold increase in plasma creatinine level and a parallel decrease in creatinine clearance that resolved when treatment with the drug was discontinued. In two thalassemic patients, diuresis was evident by urine output exceeding fluid intake. The mechanism was studied in dogs that exhibited an acute and significant decrease in inulin and para-aminohippuric acid clearances induced by intravenous deferoxamine. Saline diuresis could prevent the decrease in the glomerular filtration rate but not the decrease in renal blood flow caused by deferoxamine. Deferoxamine induced an acute increase in the fractional excretion of sodium, potassium, chloride, phosphate, and urate, which may explain the relative diuresis observed in two of the patients. In a subsequent experiment, ferrioxamine induced an increase in the fractional excretion of sodium and chloride but did not affect the glomerular filtration rate and renal blood flow. Our studies suggest that adequate hydration may be needed to preserve renal hemodynamics during intravenous deferoxamine therapy. Repeated measurements of renal function should accompany treatment with this agent.

Adolescent↗

PubMed1989

Prolongation of acute renal failure in two patients with hemolytic-uremic syndrome due to excessive plasma infusion therapy.

Two children with prototypic hemolytic-uremic syndrome had prolonged acute dialysis-dependent renal failure (74 and 84 days) associated with a state of hyperproteinemia induced by extensive infusion of fresh frozen plasma (283 and 307 units). We believe that the hyperproteinemia prolonged the duration of renal failure. Following cessation of plasma therapy, the hyperproteinemic state reversed, the degree of proteinuria decreased and renal function quickly recovered. Although the pathophysiological mechanism requires further evaluation, we speculate that an alteration in the colloid oncotic pressure and/or aggravation of tubulointerstitial injury due to overload-proteinuria may have increased the duration of renal failure.

Acute Kidney Injury↗

PubMed1989

Abnormalities in the NC1 domain of collagen type IV in GBM in canine hereditary nephritis.

Samoyed hereditary glomerulopathy (SHG) in dogs serves as a model for human X-linked hereditary nephritis (HN). We previously showed that glomerular capillaries of affected males did not stain by immunofluorescence (IF) using serum from a patient with Goodpasture's syndrome. Our goal in the present study was to determine whether the NC1 domain of the collagen type IV molecule, which contains Goodpasture antigen (GPA), could be demonstrated in these dogs, and to assess its immunological reactivity. By SDS-PAGE, NC1 in collagenase digests of glomerular basement membranes (GBM) of unaffected and carrier female dogs in the family with SHG showed 24 kilodalton (kD), 26 kD and 28 kD monomer, and 46 kD and 47 kD dimer components, but the 24 kD monomer was diminished in the affected males. By IF, a rabbit antibody to NCl stained glomerular capillaries of unaffected, affected male, and carrier female dogs. In contrast, a human anti-GBM plasmapheresis fluid (PPF) stained glomerular capillaries of only the unaffected and carrier female dogs. By RIA, both antibodies reacted strongly with NCl in collagenase digests of GBM of the unaffected and carrier female dogs, but showed reduced reactivity with NCl of affected males. By Western blotting, both antibodies bound to dimers and 24 kD and 26 kD monomers of the NCl domain in collagenase digests of GBM of unaffected and carrier female dogs. However, in affected males, the rabbit anti-NCl antibody did not bind to the 24 kD monomer, while the human anti-GBM PPF showed weak binding to the 24 kD and 26 kD monomers.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1989

A novel maturation marker for human Sertoli cells.

The maturation of Sertoli cells at puberty is critical for the initiation and maintenance of spermatogenesis. Little is known about the factors which control Sertoli cell maturation. We have examined Sertoli cells in testicular specimens from 44 subjects, ranging in age from 6 months to 67 years, for reactivity with a mouse monoclonal antibody (M2A) using the immunoperoxidase reaction. We found positive reactivity with prepubertal but not postpubertal Sertoli cells, suggesting that the antigen defined by M2A was a marker of immature Sertoli cells. This marker may be useful for studying the factors which influence Sertoli cell maturation during development of the testis.

Adolescent↗

PubMed1989

Characterization of the NC1 domain of collagen type IV in glomerular basement membranes (GBM) and of antibodies to GBM in a patient with anti-GBM nephritis.

Anti-glomerular basement membrane (GBM) nephritis is associated with production of antibodies to the Goodpasture antigen (GPA) component of the NC1 domain of collagen type IV. We studied a patient with anti-GBM nephritis with regard to 1) reactivity of the anti-GBM antibodies in his serum, plasmapheresis fluid (PPF), and an eluate prepared from GBM of his nephrectomy specimen, and 2) electrophoretic and immunologic properties of the NC1 domain extracted by collagenase digestion from GBM of his nephrectomy specimen. Antibodies to different NC1 determinants in serum, PPF and eluate were detected by immunofluorescence of glomerular capillaries of normal kidney. In addition, the antibody in PPF, but not in the eluate, reacted strongly in a plate-binding radioimmunoassay with NC1 domain extracted from normal human GBM, and bound by Western blotting to both dimer (46 kD and 49 kD) and monomer (24 kD, 26 kD and 28 kD) components of the NC1 domain. Analysis of the NC1 domain in the patient's GBM by SDS-PAGE showed a number of abnormalities, including an absence of monomer bands. Moreover, there was diminished reactivity of the patient's NC1 domain by the radioimmunoassay and Western blotting, using his PPF and a rabbit anti-NC1 antiserum. These findings indicated that there were different types of antibodies to NC1 domain in PPF and eluate, associated with an abnormal NC1 domain in GBM. These results have allowed us to speculate on the pathogenesis of anti-GBM nephritis in this patient.

Anti-Glomerular Basement Membrane Disease↗

PubMed1989

The NC1 domain of collagen type IV in neonatal dog glomerular basement membranes. Significance in Samoyed hereditary glomerulopathy.

Patients with hereditary nephritis (HN) present with renal disease after infancy, suggesting that the lesion of glomerular basement membranes (GBM) may not be congenital. Therefore, the NC1 domain of collagen type IV in normal neonatal dog GBM was compared with NC1 in normal adult GBM by SDS-PAGE and Western blotting, using two anti-NC1 antibodies. Similar results were obtained, indicating that the NC1 domain is present and immunoreactive in the neonatal period. Next, serial renal biopsies were performed on a family of Samoyed dogs with hereditary glomerulopathy (SHG), an animal model of HN, and assessed by immunofluorescence. One of the anti-NC1 antibodies produced global staining of GBM in unaffected dogs, and global/segmental staining in carrier females; however, no staining was seen in affected males as early as the neonatal period. Electron microscopy (EM) failed to demonstrate any lesion of GBM in neonatal dogs. Thus, in SHG, and presumably in human HN, the abnormality in the NC1 domain is congenital, and precedes the changes seen by EM in GBM.

Animals↗

PubMed1988

An immunohistochemical and electron microscopic study of extra-renal basement membranes in dogs with Samoyed hereditary glomerulopathy.

Samoyed hereditary glomerulopathy (SHG) in dogs has been employed as a model for human hereditary nephritis (HN), since affected dogs and patients show splitting of glomerular capillary basement membranes by electron microscopy (EM) and absent staining of glomerular capillaries for Goodpasture antigen (GPA) by immunofluorescence (IF). EM and IF were used to examine basement membranes (BM) in skin, lung, choroid plexus, lens, retina, and inner ear in SHG. By EM, BM in these tissues appeared similar in affected male, carrier female, and unaffected dogs. By IF, GPA could be detected only in lens capsule, internal limiting membrane of retina and basilar membrane of inner ear of unaffected and carrier female dogs, but not in affected male dogs. However, eye abnormalities and hearing loss were not present in any dogs, in contrast to their frequent occurrence in human HN. Our findings on extra-renal BM in SHG suggest that GPA is not required to maintain normal vision or hearing in affected male dogs and permit a greater understanding of the pathogenesis of human HN.

Animals↗

PubMed1988

Phenotypic and molecular characterization of inducible human neuroblastoma cell lines.

Two new neuroblastoma (NB) cell lines, NUB-6 and NUB-7, were established from recurrent and primary NB tumours respectively and identified conclusively as NB by their phenotypic characteristics, catecholamine production and N-myc amplification. The cell lines could be distinguished on the bases of distinctive growth patterns in monolayer culture and semi-solid media (collagen gel and agarose), neurite formation and their response to four classes of growth and differentiation modulators. The NUB-6 cell line consisted of two distinct cell subtypes, small typical neuroblasts and larger spheroid-forming cells, while NUB-7 was homogeneously neuroblastic. Class-I agents (dibutyrl cyclic AMP [dbcAMP], butyrate, and papaverine) inhibited growth of both cell lines, while only dbcAMP stimulated the formation of short neurites by NUB-6 neuroblast cells in monolayer culture and collagen. Of the class-II agents (vitamins), retinoic acid inhibited growth of both cell lines and stimulated formation of long neurites by NUB-6 cells and NUB-7 cells in later passages. In contrast, vitamin E inhibited growth of NUB-6 and late-passage NUB-7, but stimulated early passage NUB-7. The class III agent (nerve growth factor) resembled vitamin E. The class-IV agents (interferons; rIFN-alpha 2a and rIFN-gamma 1) inhibited growth of both cell lines in monolayer culture and agarose, but stimulated NUB-6 neuroblasts and early passage NUB-7 cells to form long neurites. Thus phenotypically distinct NB cell lines were established in vitro and shown to be differentially influenced by various growth and differentiation modulators. The potent effect of IFN suggests a role for these modulators in NB behaviour in vivo.

Cell Aggregation↗

PubMed1988

A monoclonal antibody as a marker for carcinoma in situ germ cells of the human adult testis.

Carcinoma in situ of the testis (CIS) is a precursor of invasive testicular germ cell tumours. The diagnosis of CIS is however often missed when conventional histological techniques are used. No specific immunological marker for CIS germ cells of the testis has been demonstrated previously. A novel monoclonal antibody, M2A, reacting with malignant germ cells of seminomas has recently been developed. Using the immunoperoxidase reaction on tissue sections, we tested the reactivity of M2A with CIS germ cells of the human adult testis. Positive reaction was found in 19 of 20 testicular specimens showing CIS, whereas no staining was found in 39 testicular biopsies without CIS. Thus, M2A may serve as a diagnostic marker in detection of CIS germ cells.

Adult↗

PubMed1988

Use of anti-seminoma monoclonal antibody to confirm the diagnosis of mediastinal seminoma. A case report.

Chest x-ray in a 20-year-old man showed a large anterior mediastinal mass and a needle aspirate was diagnosed by light microscopy (LM) as non-Hodgkin's lymphoma. Treatment with CHOP (cyclophosphamide, adriamycin, vincristine and prednisone) was ineffective and a tissue biopsy was performed. LM showed large, non-cohesive cells with abundant cytoplasm and rounded nuclei. Differential diagnoses included malignant lymphoma, seminoma, thymoma, anaplastic carcinoma, malignant melanoma and paraganglioma. Electron microscopy was not conclusive and immunoperoxidase staining was carried out. The malignant cells were negative for common leukocyte antigen, Leu M1, alpha-fetoprotein, chorionic gonadotrophin, cytokeratin, epithelial membrane antigen, carcinoembryonic antigen, S-100 protein and neuron-specific enolase but positive for placental alkaline phosphatase. In addition, there was strong positivity with a monoclonal antibody (mAb) which was recently shown to react with testicular seminomas. This case illustrates the value of this mAb in confirming the diagnosis of mediastinal seminoma.

Adult↗

PubMed1988

Reactivity of tumor cells in malignant effusions with a panel of monoclonal and polyclonal antibodies.

A panel of 13 mouse monoclonal antibodies (mAb) and 1 rabbit polyclonal antibody was tested for reactivity with tumor cells in 26 effusions obtained from patients with carcinoma of ovary, breast, or mesothelioma, using an immunoperoxidase staining reaction. Specific staining of tumor cells but not reactive mesothelial cells was demonstrated with some of the mAb in the panel. In 4 of 26 effusions no evidence of malignancy was obtained after routine cytological staining, but this was reversed on the basis of immunoperoxidase staining of tumor cells with the mAb in the panel. Serial effusions were evaluated in 4 patients during the course of chemotherapy, allowing an assessment of effect of therapy on the antigenic characteristics of the tumor cells. In another 4 patients, the results of immunoperoxidase staining of effusions were compared with those obtained by applying the same antibody panel to solid tumor nodules. There was a tendency to develop changes in the pattern of reactivity during therapy, and the pattern of reactivity was more restricted in tumor nodules than in effusions. One of the mAb in our panel (2G3) was consistently shown to produce strong staining of a high proportion of tumor cells in effusions and tumor nodules from patients with ovarian or breast cancer and may be of value in immunocytochemical diagnosis and therapy of epithelial malignancies.

Animals↗

PubMed1988

Binding of peanut agglutinin to neuroblastomas and ganglioneuromas: a marker for differentiation of neuroblasts into ganglion cells.

Binding of peanut agglutinin (PNA), concanavalin A, Ricinus communis agglutinin, wheat germ agglutinin, soybean agglutinin, Ulex europaeus agglutinin 1, and Dolichos biflorus agglutinin was assessed in tissue sections of 4 cases of neuroblastoma (NB) to determine whether there were any differences in lectin binding among immature neuroblasts, maturing neuroblasts, and ganglion cells. Only PNA binding proved to be useful. This was assessed more fully in 17 cases of NB and 7 ganglioneuromas. PNA bound to immature neuroblasts in only 4 of 12 stroma-poor NB but to maturing neuroblasts and ganglion cells in 5 of 5 stroma-rich NB. PNA bound to ganglion cells in all 7 ganglioneuromas. The NB were categorized as favorable (7 cases) and unfavorable (10 cases), using the patients' ages and the histologic appearance and mitotic-karyorrhetic index of the resected tumor, and this was correlated with PNA binding. PNA bound to 6 of 7 NB with a favorable, and 3 of 10 with an unfavorable histologic type of NB (p less than 0.05). These studies suggest that neuroblasts acquire the capacity to bind to PNA as they differentiate into ganglion cells and that PNA binding is indicative of a favorable histologic type of NB.

Age Factors↗

PubMed1988

Characterization of glomeruli by immunohistochemistry and electron microscopy in a case of Wilms' tumor.

A patient with an abdominal mass was found to have a Wilms' tumor. Light microscopic examination showed that, in addition to blastema, stroma, and tubules, the tumor contained an unusually large number of glomeruli. Glomerular basement membranes stained positively by immunohistochemistry for laminin, collagen type IV, fibronectin, and Goodpasture antigen. Staining for Goodpasture antigen was seen only after acid-urea treatment, which was similar to findings in fetal and infant glomeruli. Glomerular cells stained positively for actin, myosin, and desmin, but there was no staining for factor VIII or Ulex europaeus agglutinin I, indicating an absence of endothelial cells. These findings were supported by electron microscopy, which showed basement membrane material, visceral epithelial cells, and mesenchymal cells (presumably primitive mesangial cells) in glomeruli, but no patent capillaries or capillary endothelium. Hence, the glomeruli in this case of Wilms' tumor were immature and also showed aberrant glomerulogenesis.

Female↗