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B Rozell

Publications and source records attributed to B Rozell.

At least 19 recordsLinked to original sources
PubMed2006

The hepatitis C virus and immune evasion: non-structural 3/4A transgenic mice are resistant to lethal tumour necrosis factor alpha mediated liver disease.

BACKGROUND: The hepatitis C virus (HCV) establishes chronic infection by incompletely understood mechanisms. The non-structural (NS) 3/4A protease/helicase has been proposed as a key complex in modulating the infected hepatocyte, although nothing is known about the effects this complex exerts in vivo. AIM: To generate mice with stable and transient hepatocyte expression of the HCV NS3/4A proteins to study its effects in vivo. METHODS: NS3/4A expression was determined by western blot and immunohistochemistry. Two independent pathologists determined the liver histology. Hepatic immunity was characterised by quantifying intrahepatic immune cell subsets. Liver damage was induced using carbon tetrachloride (CCl(4)), lipopolysaccaride (LPS), tumour necrosis factor alpha (TNFalpha), and anti-Fas antibody. RESULTS: Expression of NS3/4A was restricted to the cytoplasm of hepatocytes, and did not cause liver cancer or any spontaneous liver pathology. However, the presence of NS3/4A modulated the intrahepatic immunity, as follows: first, the CD4+ T cell and type I/II dendritic cell subsets were reduced in transgenic livers; second, NS3/4A protected hepatocytes from liver damage mediated in vivo by CCl(4), LPS, TNFalpha, but not FAS; and third, both stable and transiently NS3/4A transgenic mice were resistant to lethal doses of liver targeted TNFalpha, and the resistance could be reverted by treatment with a p38 mitogen activated protein kinase inhibitor (MAPK). CONCLUSIONS: Hepatic expression of NS3/4A does not induce spontaneous liver disease. NS3/4A does, however, alter the intrahepatic immune cell subsets and protects hepatocytes against TNFalpha induced liver damage in vivo. The TNFalpha resistance can be reverted by treatment with a p38 MAPK inhibitor. This represents a new immune evasion strategy conferred by NS3/4A.

Animals↗
PubMed2003

Low dose and gene gun immunization with a hepatitis C virus nonstructural (NS) 3 DNA-based vaccine containing NS4A inhibit NS3/4A-expressing tumors in vivo.

The hepatitis C virus (HCV) protease and helicase encompasses the nonstructural (NS) 3 protein and the cofactor NS4A, which targets the NS3/4A-complex to intracellular membranes. We here evaluate the importance of NS4A in NS3-based genetic immunogens. A full-length genotype 1 NS3/4A gene was cloned into a eucaryotic expression vector in the form of NS3/4A and NS3 alone. Transient transfections revealed that the inclusion of NS4A increased the expression levels of NS3. Subsequently, immunization with the NS3/4A gene primed 10- to 100-fold higher levels of NS3-specific antibodies as compared to immunization with the NS3 gene. Humoral responses primed by the NS3/4A gene had a higher IgG2a/IgG1 ratio (>20) as compared to the NS3 gene (3.0), suggesting a T helper 1-skewed response. Low dose i.m. (10 microg) immunization with the NS3/4A gene inhibited the growth of NS3/4A-expressing tumor cells in vivo, whereas the NS3 gene alone or NS3 protein did not. We then evaluated the efficiency of the NS3/4A gene administered by the gene gun, at the same doses used for humans, in priming cytotoxic T lymphocyte (CTL) responses. Three to four 4 microg doses of the NS3/4A gene primed CTL at a precursor frequency of 2-4%, which inhibited the growth of NS3/4A-expressing tumor cells in vivo. Thus, NS4A enhances the expression levels and immunogenicity of NS3, and an NS3/4A gene delivered transdermally could be a therapeutic vaccine candidate.

Animals↗
PubMed2000

Direct oral microscopy and its value in diagnosing mucosal lesions: a pilot study.

OBJECTIVE: Clinical examination of the oral mucosa often leads to an uncertain diagnosis, and a supplementary biopsy with a histopathologic examination of the lesion is necessary to establish a definite diagnosis. However, the site for the biopsy is a subjective choice that sometimes raises doubts about its representativeness. So far, no simple and reliable method is available for selecting the most appropriate area for biopsy. STUDY DESIGN: In a prospective study, we performed direct oral microscopy (oral application of the colposcopy technique used in gynecology) in 35 patients with various clinical diagnoses, such as leukoplakia, oral lichenoid lesions, or suspected malignancy. First, the oral mucosa was examined with direct microscopy, and the most representative site, according to colposcopic criteria, was selected. Then, the mucosa was clinically inspected by an independent examiner. The best site for biopsy according to clinical criteria was noted, and any difference in biopsy sites was recorded. Biopsy specimens were taken from 2 of these sites. RESULTS: Twenty-nine patients (83%) showed changes in the vascular picture on microscopy, according to the colposcopy criteria. In 14 patients (40%), the biopsy sites identified by direct oral microscopy showed more advanced histologic signs than those selected by routine clinical examination (0.01 < P </=.05). Four patients (11%) had more advanced histologic signs in the biopsy samples, as identified during routine clinical examination. In 17 patients (49%), we found no differences between the biopsy specimens. CONCLUSION: Direct oral microscopy of mucosal lesions seems to offer advantages in selecting more representative sites for biopsy than routine clinical examination alone.

Adult↗
PubMed1998

A conserved structural element in horse and mouse IGF2 genes binds a methylation sensitive factor.

The equine IGF2 gene has been cloned and characterised. It spans a 9 kb region, which is substantially less than the corresponding human gene. Three coding exons and three untranslated leader exons, all highly homologous to those in other species, were identified. Downstream of the polyadenylation site in exon 6, a dinucleotide repeat sequence was identified. Three putative promoters (P1-P3) were localised in the 5' region of the gene. RNase protection analysis revealed two active promoters in fetal tissues, P2 and P3, whereas P3 was the only promoter active in adult tissues. This represents a transcriptional pattern different from that in humans or rodents. A novel structural element, an inverted repeat, is predicted in the 3' region of the IGF2 gene. This repeat is conserved between species and located in a region which is differentially methylated in the human and mouse genes and might therefore be involved in the imprinting mechanism. The inverted repeat acquires a stem-loop structure in vitro with a hybrid A/B-DNA conformation in the stem area. Both in horse and mouse, a methylation-sensitive protein binds this structure with a strong requirement for the loop area. Furthermore, the protein might be developmentally regulated.

Animals↗
PubMed1998

Dynamic expression of E-cadherin in ameloblasts and cementoblasts in mice.

During embryonic development, E-cadherin mediates intercellular adhesion in a variety of epithelia in a spatio-temporal pattern. We have analyzed the distribution of this cell adhesion molecule in the mouse during odontogenesis, at both mRNA and protein levels, in the mandibular first molars and incisors. E-cadherin was strongly expressed at the bell stage by the cells of the dental organ, and by the pre-secretory ameloblasts and the cells of stratum intermedium at the early mineralization stage. At the onset of enamel secretion, E-cadherin disappeared from the apical pool of the ameloblasts and was later absent from the post-secretory ameloblasts. E-cadherin was also found in Hertwig's root sheath and later in the cells producing acellular cementum. These findings indicate that E-cadherin may be involved in the polarization of the ameloblasts and in the early stages of cementogenesis.

Ameloblasts↗
PubMed1998

Expression of the insulin-like growth factor II gene in polychlorinated biphenyl exposed female mink (Mustela vison) and their fetuses.

AIM: To study how polychlorinated biphenyls (PCBs) affect fetal growth and the expression of the insulin-like growth factor (IGF II) gene in the mink (Mustela vision). METHODS: Ten female mink were each exposed to 0.65 or 1.3 mg Clophen A50/day, respectively, during the reproductive season. The numbers and sizes of viable fetuses were recorded. The expression of the IGF II gene was studied by northern blotting using a mink specific IGF II cDNA probe. RESULTS: The number of viable fetuses decreased after PCB exposure in a dose dependent fashion. Expression of the IGF II gene in adult livers from PCB exposed animals was decreased, compared with control animals, in a dose dependent fashion. In contrast, IGF II expression in placentas and fetuses was unaltered. Furthermore, the maternal excretion of urinary cortisol increased in both exposed groups during the implantation period. CONCLUSIONS: Expression of the IGF II gene is downregulated by PCB exposure in the adult liver. There is also an indication that the regulation of the expression of this gene differs between adult and fetal life.

Animals↗
PubMed1997

FISH mapping of the IGF2 gene in horse and donkey-detection of homoeology with HSA11.

Three genomic subclones derived from a phage clone containing the equine IGF2 gene were used to FISH map the gene on horse (ECA) and donkey (EAS) metaphase chromosomes. The gene mapped on ECA 12q13 band and is the first locus mapped to this horse chromosome. In donkey the gene mapped very terminal on the long arm of one small submetacentric chromosome that shows almost identical DAPI-banding pattern with ECA12. This is the first locus mapped in donkey genome. Cross species chromosome painting of equine metaphase chromosomes with human Chromosome (Chr) 11-specific probe showed homoeology of this human chromosome with ECA12 and ECA7. The novel ECA12 comparative painting results are thus in accordance with the localization of the equine IGF2 gene. Comparison of the hitherto known physical locations of IGF2 in different species, viz. human, cattle, sheep, horse, and donkey, shows that this gene tends to maintain a terminal location on the chromosome arm.

Animals↗
PubMed1997

Neurological disease and encephalitis in cats experimentally infected with Borna disease virus.

Barrier-bred cats were inoculated intracerebrally with either the rabbit-adapted Borna disease virus (BDV) strain V or a newly isolated feline BDV, obtained from a cat with natural staggering disease (SD). Three out of eight inoculated cats developed neurological signs and non-suppurative encephalitis; all three recovered from the acute stage of disease. Sero-conversion and the development of neutralizing antibodies occurred in all of the virus-inoculated cats. In addition, cats inoculated with feline BDV showed an early peripheral T cell response not present in cats inoculated with BDV strain V, suggesting that the feline virus exerted a more vigorous effect on the immune system. Using immunohistochemistry and a reverse transcriptase-polymerase chain reaction assay, BDV-specific antigen and nucleic acid could be demonstrated in brain samples from each cat with encephalitis, showing that incomplete viral clearance was probably responsible for the maintenance of inflammation. The successful induction of neurological signs and encephalitis in one cat infected with feline BDV, together with the detection of BDV-specific antigen and nucleic acid in the brain, provides strong evidence for the notion that BDV is the etiological agent behind feline SD.

Animals↗
PubMed1996

Cloning and sequencing of an equine insulin-like growth factor I cDNA and its expression in fetal and adult tissues.

A cDNA for equine insulin-like growth factor I (IGF I) has been isolated by reverse transcriptase-polymerase chain reaction and subsequently sequenced. The sequenced fragment contained 465 bp including the coding regions for the signal peptide, the entire mature protein, and 4 amino acids into the E-peptide. Like its human counterpart, the mature equine IGF I peptide contains 70 amino acids and was 100% homologous between horse and man. The 49-amino-acid signal peptide had the threonine in position 26 of the human signal peptide substituted by isoleucine. The nucleotide homology across the entire clone was 96.3% between horse and man and 91.6% between horse and rat. The isolated cDNA hybridized to the same transcripts in fetal and adult tissues.

Amino Acid Sequence↗
PubMed1995

C-CAM expression in odontogenesis and tooth eruption.

The distribution of the cell adhesion molecule C-CAM was analyzed during tooth development, eruption and formation of the junctional epithelium in rat molars by immunohistochemistry and in situ hybridization. C-CAM could not be detected during odontogenesis until the late bell stage; then only the mRNA was demonstrated in the odontoblasts and ameloblasts. Prior to eruption, a local increase in C-CAM (mRNA and protein) was observed in the reduced enamel epithelium. During eruption, high C-CAM levels were seen in the fusion zone between the oral epithelium and the reduced enamel epithelium. In the adult rat, C-CAM remained strongly expressed in the junctional epithelium. Our study indicates that C-CAM may play a role in odontogenesis and during formation of the epithelial structures involved in tooth eruption and formation of the junctional epithelium.

Ameloblasts↗
PubMed1994

Histologic changes in dental morphology induced by high dose chemotherapy and total body irradiation.

Disturbances in dental development were studied with the use of radiography and histology in a patient with acute lymphoblastic leukemia who was treated with induction chemotherapy at 2.3 years of age and bone marrow transplantation at 4.3 years of age. The follow-up 9.5 years after bone marrow transplantation showed evidence of short tapered roots, enamel hypoplasia, microdontia, and aplasia. A histologic examination of two extracted permanent teeth showed that the crown of the maxillary lateral incisor exhibited numerous incremental lines that corresponded closely to the treatment periods with cytotoxic drugs. The maxillary second premolar exhibited regularly spaced incremental lines in the enamel and dentine. A gross hypoplasia was seen in the cervical part of the crown corresponding to the time of administration of 10 Gy total body irradiation. The results indicate that chemotherapy mainly induces qualitative disturbances in dentine and enamel, whereas total body irradiation induces both qualitative and quantitative changes.

Abnormalities, Radiation-Induced↗
PubMed1994

Distribution of protein kinase C alpha and accumulation of extracellular Ca2+ during early dentin and enamel formation.

Activation of the protein kinase C (PKC)-related signal transduction system has been associated with phenotypic expression in a wide variety of cell types. In in vitro studies, it has often been activated by relatively small increases in the Ca2+ concentration ([Ca2+]) in the medium. The studies reported here explored the hypothesis that localized increases in the extracellular [Ca2+] and activation of the PKC-related pathway may be involved in early dentin and enamel formation. Whole-head, freeze-dried sections through the developing molars of 5-day-old rats were evaluated by methods that localized non-crystalline Ca2+. Immunohistochemical methods were adapted for use with the freeze-dried sections, and two monoclonal antibodies were used to localize PKC alpha in the formative cells of the developing teeth. Low concentrations of extracellular Ca2+ were observed in the early, unmineralized dentin in the area of ameloblast differentiation. Increased concentrations occurred at the point of initial dentin mineralization, immediately before the beginning of enamel matrix deposition. PKC alpha was localized in the differentiating odontoblasts, at the beginning of dentin matrix deposition. It was intensely localized in the distal borders of the pre-ameloblasts, and appeared to redistribute in the cells during ameloblast differentiation. These observations suggest that local increases in the extracellular [Ca2+] and the PKC signal transduction pathway may be involved in key inductions in the early stages of dentin and enamel formation.

Ameloblasts↗
PubMed1993

Glutathione transferases of classes alpha, mu and pi show selective expression in different regions of rat kidney.

1. Glutathione transferases (GST) are mainly cytosolic and occur in multiple forms, which can be arranged in three distinct, structural classes. The different enzyme forms show distinct substrate specificities with electrophilic and genotoxic substances. The expression of the alpha subunits 1, 2 and 8, the mu subunits 3, 4 and 6, and the pi subunit 7 of GST in different parts of the rat kidney was determined immunohistochemically. 2. GST immunoreactivity was present predominantly in the nephron, collecting duct and urothelium. 3. A conspicuous finding was that subunits 1, 2 and 8 were localized to the proximal tubules, while the mu subunit 3 was demonstrable in epithelial tubular cells from the distal tubules to the urothelium. The immunoreactivity of subunits 4 and 6 could be visualized in epithelial cells from the ascending thin limb to the collecting ducts. Subunit 7 was found in the thin limb of the loop of Henle, and in scattered cells in the distal tubules. 4. The urothelial cells covering the papilla and the renal calyces showed immunoreactivity to GST subunits 2-4 and 6-8. 5. Thus, in the nephron the class alpha GSTs were selectively expressed in the proximal tubules and the class mu and class pi GST in the thin loop of Henle and distal tubules. The cells in the collecting ducts and the urothelium, which have a different ontogeny than the nephron, do not show any corresponding differential distribution of the GST classes. 6. Cells in a given location were in some cases found to be non-reactive with a given antiserum in an otherwise immunoreactive cell population, demonstrating a spatial variation in GST expression. The immunoreactivity to the different forms of GST was predominantly cytoplasmic but a nuclear localization could also be demonstrated. 7. The panel of antibodies to GST may tentatively be used as markers in localizing lesions in restricted parts of the nephrons and to elucidate dynamic alterations in the tubular system in response to physiological and toxic agents.

Animals↗
PubMed1993

Immunochemical characterization and tissue distribution of glutaredoxin (thioltransferase) from calf.

Glutaredoxin catalyzes glutathione-dependent disulfide oxidoreduction reactions in a coupled system with NADPH, GSH and glutathione reductase and has an active site disulfide/dithiol with the sequence -Cys-Pro-Tyr-Cys-. Calf thymus glutaredoxin (thioltransferase), which contains two additional structural half-cystine residues, was purified to homogeneity, using a modification of the previously described isolation procedure. This method involved a pI-shift of glutaredoxin, obtained after oxidation of the fully reduced form with hydroxyethyl-disulfide, followed by CM-Sepharose chromatography. On both SDS- and IEF-gels the protein migrated as one band (M(r) 12,000). The pure protein was used to affinity-purify rabbit antiglutaredoxin antibodies obtained by immunization with the oxidized form of glutaredoxin. Using these antibodies the distribution of glutaredoxin was mapped in calf organs and tissues by Western blots and by immunohistochemistry. Glutaredoxin was demonstrated in all organs investigated. Western blots showed the presence of weak additional high molecular weight bands of unknown identity in certain organs. The immunohistochemical analyses revealed that glutaredoxin is highly expressed in a wide variety of cell types, both epithelial and mesenchymal. The distribution and occurrence in the calf organs was similar to that previously described for thioredoxin in the rat. There were some exceptions: e.g., follicular cells in the ovary did not contain immunohistochemically demonstrable glutaredoxin but expressed thioredoxin. Particularly striking were observations of strong glutaredoxin immunoreactivity in oocytes in the ovary and the pattern of glutaredoxin in epithelial tissue of the skin and tongue reflecting differential expression during cell differentiation. The distribution demonstrated that glutaredoxin serves functions apart from the originally described role as hydrogen donor for ribonucleotide reductase which only occurs in replicating cells. Such functions should relate particularly to glutathione-catalyzed protein disulfide oxidoreductions and cellular signalling by redox regulating mechanisms.

Amino Acid Sequence↗
PubMed1989

Changes in the distribution of insulin-like growth factor I, thioredoxin, thioredoxin reductase and ribonucleotide reductase during the development of the retina.

The postnatal development of the rat retina offers good opportunities to follow migration and differentiation of neuroectodermal cells. In this study we have analyzed, by immunohistochemical methods, the distribution of several protein antigens and the trophic peptide insulin-like growth factor I (IGF-I; somatomedin-C). The latter contains disulfide bonds, which in vitro are reduced by the thioredoxin system, i.e. thioredoxin, thioredoxin reductase and NADPH. Ribonucleotide reductase provides growing cells with deoxyribonucleotides, necessary for DNA synthesis, and thioredoxin is an in vitro hydrogen donor. By immunofluorescence IGF-I immunoreactivity was observed throughout the Müller neuroglial cells in the developing retina, but only to a very small extent in the mature retina. Nerve cells showed transient expression of IGF-I during their development. The IGF-I immunoreactivity is likely to be due to local synthesis, since we could demonstrate retinal IGF-I mRNA. Treatment with the transport-blocking agent colchicine caused the Müller glial cells and the retinal pigment epithelium cells to become IGF-I immunoreactive. Thioredoxin and thioredoxin reductase immunoreactivities are confined to neurons and photoreceptor cells in the developing retina. Ribonucleotide reductase subunit M1 immunoreactivity was only observed during the first postnatal week in proliferating neuroectodermal cells. In conclusion, IGF-I and ribonucleotide reductase subunit M1 immunoreactivities are transiently expressed by neuroectodermal cells in the developing rat retina. In contrast, the proteins of the thioredoxin system are demonstrable also in the mature retina.

Animals↗
PubMed1988

Immunocytochemical evidence for the cytoplasmic localization and differential expression during the cell cycle of the M1 and M2 subunits of mammalian ribonucleotide reductase.

Mammalian ribonucleotide reductase consists of two non-identical subunits, proteins M1 and M2. We have produced and characterized rat polyclonal and monoclonal antibodies directed against protein M2 of mouse ribonucleotide reductase. Using these antibodies for immunocytochemical studies, an exclusively cytoplasmic localization of protein M2 was demonstrated both in cultured parent and hydroxyurea-resistant, M2-over-producing mouse TA3 cells, and in cells from various mouse tissues. These data, together with the previously demonstrated cytoplasmic localization of the M1 subunit, clearly show that ribonucleotide reductase is a cytoplasmic enzyme. Combining the anti-M2 antibodies with a monoclonal anti-M1 antibody allowed for double-labelling immunofluorescence studies of the two subunits in individual cells. Only approximately 50% of the cells in a logarithmically growing culture contained immunodetectable protein M2, while the M1-specific staining was present in all cells. The M2 staining correlates well with the proportion of cells in the S-phase of the cell cycle. In tissues, only actively dividing cells stained with either antibody and there were always fewer cells stained with the M2-antibodies than with the M1-antibody. Our data therefore present independent evidence for the earlier proposed model of a differential regulation during the cell cycle of the M1 and M2 subunits of ribonucleotide reductase.

Animals↗
PubMed1988

Immunohistochemical localization of insulin-like growth factor I in the adult rat.

Rabbit antisera against native human insulin-like growth factor I (IGF-I; somatomedin C) or a synthetic tetradecapeptide, representing the carboxyterminal amino acids 57-70 of human IGF-I, were used to map immunohistochemically the distribution of IGF-I immunoreactive material in adult rats. Both antisera were specific for IGF-I, as characterized by immunoabsorption, immunoblotting and radioimmunoassay. There was no cross-reactivity to IGF-II, relaxin or pro-insulin; substances having a high degree of structural homology with IGF-I. High IGF-I immunoreactivity was observed in spermatocytes of the testis; in oocytes, granulosa and theca interna cells of the ovary during early stages of follicle development; in some lymphocytes and in reticular cells of lymphoid and hematopoietic organs; in salivary gland duct cells; in the adrenal medulla, the parathyroid gland and the Langerhans' islets. Chondrocytes in the epiphyseal and rib growth plates and at articular surfaces showed strong IGF-I immunoreactivity. Brown but not white fat cells were stained. Nerve cells in the peripheral and autonomic nervous system showed faint to intense IGF-I immunoreactivity. In contrast, neurons and neuroglial cells in the central nervous system were generally negative; motor neurons being an exception. Erythropoietic, thrombocytopoietic and myeloic cells in the bone marrow showed IGF-I immunoreactivity, but only at defined developmental stages. Hepatocytes showed faint IGF-I immunoreactivity, but became more intensely stained after pretreatment with colchicine. The present results suggest that IGF-I is synthetized by cells in several tissues and organs in the adult rat. There was an apparent association between the localization of IGF-I and cell differentiation.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗