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Biomedical subjects

S Karlsson

Publications and source records attributed to S Karlsson.

At least 163 records · Page 9Linked to original sources
PubMed1994

Correction of the enzyme deficiency in hematopoietic cells of Gaucher patients using a clinically acceptable retroviral supernatant transduction protocol.

Gaucher disease is a lysosomal storage disorder caused by a deficiency of the enzyme glucocerebrosidase (GC), and is an excellent candidate for gene replacement therapy. To develop a clinically acceptable protocol for this purpose, we created two amplified (A) high-titer retroviral vector-producer cell lines to efficiently transduce hematopoietic stem and progenitor cells. GP+envAm12/A-LGSN (A-LGSN), contained the GC cDNA driven by the retroviral long terminal repeat (LTR) and the neomycin phosphotransferase gene expressed from the simian virus 40 early promoter. GP+envAm12/A-LG4 (A-LG4) contained only the GC gene driven by the LTR. Both A-LGSN and A-LG4 contained multiple proviral copies and gave approximately 10-fold higher titers on 3T3 cells compared to their unamplified counterparts. These vectors were packaged in GP+envAm12 cells because vectors produced in this cell line transduced hematopoietic cells more efficiently than other packaging cells tested. Bone marrow mononuclear cells and purified CD34+ cells were infected with virus supernatants four times in the presence of interleukin-3 (IL-3), IL-6, and stem cell factor (SCF) over 96 hours in culture. Cells were then plated in semisolid cultures and colony-forming unit-granulocyte/macrophage (CFU-GM) colonies were scored for vector presence by polymerase chain reaction (PCR). Transduction efficiency of CFU-GM colonies derived from CD34+ cells was improved considerably using the amplified vectors in the GP+envAm12 packaging line. For A-LGSN, A-LG4, and unamplified LGSN, transduction efficiencies were 41, 42, and 25%, respectively. Therefore, multiple proviral copies resulting in higher titer improves retroviral transduction of human hematopoietic progenitor cells. Hematopoietic cells from Gaucher patients were transduced and placed into long-term bone marrow culture (LTBMC). Viral supernatant from the amplified producer lines transduced long-term culture initiating cells (LTCIC) efficiently (30 to 50%) using this clinically acceptable protocol. Both sustained mRNA expression and GC enzyme production are achieved in the long-term culture of LTCIC and lead to correction of the GC deficiency in their progeny cells.

Bone Marrow Cells↗
PubMed1993

Transforming growth factor beta 1 (TGF-beta 1) controls expression of major histocompatibility genes in the postnatal mouse: aberrant histocompatibility antigen expression in the pathogenesis of the TGF-beta 1 null mouse phenotype.

The phenotype of the transforming growth factor beta 1 (TGF-beta 1) null mouse has been previously described and is characterized by inflammatory infiltrates in multiple organs leading to a wasting syndrome and death as early as 3 weeks after birth. Since this phenotype occurs in the absence of any detectable pathogen, potential autoimmune disease mechanisms were investigated. We examined major histocompatibility complex (MHC) mRNA expression in tissues of the TGF-beta 1 null mouse and found levels of both the class I and class II MHC mRNA elevated compared to normal or TGF-beta 1 heterozygous littermates. This elevated expression was seen prior to any evidence of inflammatory infiltrates, suggesting a causal relationship between increased MHC expression and activation of immune cell populations. Cell surface expression of MHC molecules was detected by immunohistochemistry and correlated well with mRNA levels. Expression of mRNA for interferon gamma and its receptor was unchanged at the ages when increased MHC expression became apparent. Down-regulation of class I MHC expression by TGF-beta 1 was also demonstrated in vitro in fibroblasts isolated from TGF-beta 1 null mice. These findings suggest that one natural function of TGF-beta 1 is to control expression of both MHC classes. Altered regulation of MHC expression may be a critical step leading to the multifocal inflammation and wasting syndrome seen in the TGF-beta 1 null mouse. These results suggest potential applications for TGF-beta in the management of autoimmune disease, allograft rejection, and other problems associated with altered MHC expression.

Animals↗
PubMed1993

Loss of expression of transforming growth factor beta in skin and skin tumors is associated with hyperproliferation and a high risk for malignant conversion.

Mouse skin carcinomas arise from a small subpopulation of benign papillomas with an increased risk of malignant conversion. These papillomas arise with limited stimulation by tumor promoters, appear rapidly, and do not regress, suggesting that they differ in growth properties from the majority of benign tumors. The transforming growth factor beta (TGF-beta) proteins are expressed in the epidermis and are growth inhibitors for mouse keratinocytes in vitro; altered TGF-beta expression could influence the growth properties of high-risk papillomas. Normal epidermis, tumor promoter-treated epidermis, and skin papillomas at low risk for malignant conversion express TGF-beta 1 in the basal cell compartment and TGF-beta 2 in the suprabasal strata. In low-risk tumors, 90% of the proliferating cells are confined to the basal compartment. In contrast, the majority of high-risk papillomas are devoid of both TGF-beta 1 and TGF-beta 2 as soon as they arise; these tumors have up to 40% of the proliferating cells in the suprabasal layers. Squamous cell carcinomas are also devoid of TGF-beta, suggesting that they arise from the TGF-beta-deficient high-risk papillomas. In some high-risk papillomas, TGF-beta 1 loss can occur first and correlates with basal cell hyperproliferation, while TGF-beta 2 loss correlates with suprabasal hyperproliferation. Similarly, TGF-beta 1-null transgenic mice, which express wild-type levels of TGF-beta 2 in epidermis but no TGF-beta 1 in the basal layer, have a hyperproliferative basal cell layer without suprabasal proliferation. In tumors, loss of TGF-beta is controlled at the posttranscriptional level and is associated with expression of keratin 13, a documented marker of malignant progression. These results show that TGF-beta expression and function are compartmentalized in epidermis and epidermal tumors and that loss of TGF-beta is an early, biologically relevant risk factor for malignant progression.

Animals↗
PubMed1993

Role of extracellular Na+ on CCK-8-induced insulin secretion.

CCK-8 stimulates insulin secretion by an effect involving phosphoinositide (PI) hydrolysis and release of Ca2+ from intracellular stores. In this study, we examined the dependence of Na+ for the effects of CCK-8. CCK-8 (100 nM) stimulated insulin secretion from isolated rat islets. A first phase, which lasted 10 min, was not affected by removal of external Na+, whereas a second phase was abolished. CCK-8-stimulated 45Ca2+ and 3H efflux in 45Ca(2+)- and myo-[2-3H]-inositol-prelabelled islets were not affected by removal of external Na+. In a second series of experiments, pancreatic rat islet cells were loaded with the Ca2+ fluorophore FURA 2-AM. CCK-8 (100 nM) induced a rapid and transient increase in the cytoplasmic free Ca2+ concentration ([Ca2+]IC), followed by a subsequent reduction of [Ca2+]IC below the prestimulatory levels. The CCK-8-induced increase in [Ca2+]IC was not dependent on extracellular Na+, whereas the decline in [Ca2+]IC after the initial peak was slower in the absence than in the presence of Na+. Thus, the present study shows that both the second phase of CCK-8-stimulated insulin secretion and the CCK-8-stimulated postpeak reduction in [Ca2+]IC are dependent on extracellular Na+.

Animals↗
PubMed1993

Transforming growth factor beta 1 null mutation in mice causes excessive inflammatory response and early death.

To delineate specific developmental roles of transforming growth factor beta 1 (TGF-beta 1) we have disrupted its cognate gene in mouse embryonic stem cells by homologous recombination to generate TGF-beta 1 null mice. These mice do not produce detectable amounts of either TGF-beta 1 RNA or protein. After normal growth for the first 2 weeks they develop a rapid wasting syndrome and die by 3-4 weeks of age. Pathological examination revealed an excessive inflammatory response with massive infiltration of lymphocytes and macrophages in many organs, but primarily in heart and lungs. Many lesions resembled those found in autoimmune disorders, graft-vs.-host disease, or certain viral diseases. This phenotype suggests a prominent role for TGF-beta 1 in homeostatic regulation of immune cell proliferation and extravasation into tissues.

Alleles↗
PubMed1993

Evaluation of expression of transferred genes in differentiating myeloid cells: expression of human glucocerebrosidase in murine macrophages.

The retroviral vector LGSN, in which the human glucocerebrosidase (GC) cDNA is driven by the Moloney murine leukemia virus (MoMLV) long terminal repeat (LTR), was tested for expression in the murine myelomonocytic leukemia cell line M1 before and after induction of differentiation with interleukin-6 (IL-6). Southern analysis of the seven transduced clones selected for neomycin resistance in Geneticin (G-418 sulfate) demonstrated one to eight copies of intact provirus with rearrangements in only two clones. Absolute levels of human GC RNA and protein increased with increased copy numbers of provirus in the clones. Upon induction with IL-6 of the seven transduced clones to the macrophage phenotype, there was no significant change, overall, in RNA levels but some increase in human GC protein levels could be detected. Although this was the average trend, considerable clonal variation in RNA and protein levels was observed upon induction. Transduction of the M1 cells did not interfere with the ability of the cells to differentiate from blasts to macrophages as seen by the appearance of membrane receptors for the constant region of immunoglobulins (Fc gamma RI) and lysozyme production in the differentiated M1 cells. Thus, the M1 cell line can be used for testing retroviral vector expression in myeloid lineages at early and late stages of differentiation. This rapid in vitro testing of potential retroviral vectors will be beneficial for gene therapy of disorders that affect differentiated macrophages such as Gaucher's disease.

Animals↗
PubMed1993

Idiopathic cervical lesions: in vivo investigation by oral microendoscopy and scanning electron microscopy. A pilot study.

The development and progression of cervical lesions have not been thoroughly documented. Clinically, two morphologically distinct forms, saucer-or wedge-shaped, have been described. In this study, the micromorphology of manifest cervical lesions was documented in six subjects, by SEM replication and microendoscopy. Optoelectronic recordings were made of extreme mandibular lateral excursions. Salivary parameters were evaluated and a detailed case history, including oral hygiene and dietary habits, was taken. Lesions of varying severity could be observed in the same subject. Longitudinal fractures of the enamel were common. The dietary analyses were uneventful and salivary values were normal. No correlation was found between brushing habits and the location and severity of the lesions. The optoelectronic recordings, however, indicated a possible correlation between irregular lateral excursion and the severity and location of the lesions, with a tendency for fewer and less severe lesions on the preferred chewing side. The results have indicated some areas of interest for future studies applying non-invasive observation methods.

Adult↗
PubMed1993

Speech motor control in Parkinson's disease: a comparison between a clinical assessment protocol and a quantitative analysis of mandibular movements.

Nine individuals with Parkinson's disease (PD) and nine normal control subjects, matched for sex and age, were compared on a standardized assessment protocol for dysarthria. Simple measures of hand and gait motility were also obtained. A kinematic analysis of vertical jaw movements during speech was performed by using an optoelectronic technique. The PD subjects were examined during both induced off L-dopa and on L-dopa states, and their performance was compared between the two conditions. Significant group differences were found for all kinematic measures during a syllable repetition task, as well as for the total dysarthria test scores and certain individual test items. Implications and clinical usefulness of the kinematic approach to the study of oral motor functions are discussed.

Aged↗
PubMed1993

The fit of gold inlays and three ceramic inlay systems. A clinical and in vitro study.

Four inlay systems--gold, Cerec, Mirage, and Empress inlays--were evaluated for their adaptation to stone die and clinically to the tooth by means of a replica technique. Twenty inlays of each system were placed on premolars and molars in the lower jaw. A microscope was used to measure the adaptation at the approximal margin, at the inner axial wall, and at the occlusal cavosurface area. An overall better fit was observed for the gold inlays than for the ceramic inlays. When the different measuring locations were compared, a better fit was found for the occlusal area. The greatest discrepancies were recorded for the Cerec inlays, whereas the Mirage and Empress inlays were comparable.

Adult↗
PubMed1993

The fit of Procera titanium crowns. An in vitro and clinical study.

In contrast to conventional casting methods for the production of a metallic crown the Procera process is based on precision machine duplication of models combined with spark erosion of the metal frame. Some of the steps in the manual handling procedure are thereby excluded. The present study evaluated, by means of a replica technique, the fit or adaptation of Procera titanium crowns to the stone die and in vivo to the tooth before cementation. For any combination, the marginal adaptation was superior to and significantly better than the occlusal areas and axial surfaces, respectively. In addition, the crowns had a significantly better fit to the stone die than to the tooth. The marginal discrepancy was approximately 60 microns in vitro and 70 microns in vivo, with a range of 3-205 microns.

Crowns↗
PubMed1993

Occlusal force and mandibular movements in patients with removable overdentures and fixed prostheses supported by implants in the maxilla.

Thirteen patients with complete dentures in the maxilla were treated with implants to support a removable bar-retained overdenture for 1 year. Thereafter, the patients were provided with fixed prostheses and were followed up for another year. Maximal occlusal force and mandibular movements were recorded with and without the bar supporting the denture and then during the subsequent year with fixed prostheses. Significant changes in occlusal force capacity and mandibular movements were observed when the bar was removed. Less obvious functional changes were found when the fixed prostheses were placed, with the exception of occlusal force levels, which increased significantly over time.

Aged↗
PubMed1993

Gene transfer and bone marrow transplantation with special reference to Gaucher's disease.

Gene therapy following gene transfer into hematopoietic cells is now being investigated for several genetic disorders (1). The candidate diseases under consideration and active experimental investigation are disorders that can be fully or partially corrected by allogeneic bone marrow transplantation, where the gene defect is known and the normal gene has been cloned and characterized. Together with ADA deficiency Gaucher's disease is a leading candidate among these disorders. We have developed high titer retroviral vectors containing the glucocerebrosidase gene that can transduce murine bone marrow stem cells with high efficiency and will result in high levels of human glucocerebrosidase in macrophages of long-term reconstituted mice. "Gene therapy" for Gaucher's disease has therefore been accomplished in the mouse. We have also developed high titer amphotropic vectors containing the glucocerebrosidase gene that can transduce human CD34+ hematopoietic progenitors with a high degree of efficiency. Up to 70% of CFU-GM colonies from these infected CD34+ cells contained the transferred gene as determined by PCR. Our vectors can also correct the enzyme deficiency of hematopoietic cells from Gaucher patients following gene transfer.

Animals↗
PubMed1992

High levels of human glucocerebrosidase activity in macrophages of long-term reconstituted mice after retroviral infection of hematopoietic stem cells.

Gaucher disease is a leading candidate for somatic gene therapy using bone marrow (BM) cells as target tissue. Towards this end, we have constructed a retroviral vector (LG) in which the human glucocerebrosidase (GC) cDNA is driven by the Moloney murine leukemia virus (MoMLV) long terminal repeat (LTR). Day 12 to 14 colony-forming unit-spleen progenitor cells were infected by the LG virus with a 100% efficiency, and GC messenger RNA (mRNA) and protein were detected in the progeny of these cells. Tissues from long-term reconstituted mice analyzed 8 months posttransplantation with LG-infected BM contained the intact provirus at greater than 1 copy per cell, indicating effective infection of hematopoietic stem cells. Human GC mRNA generated by the viral LTR was detected in macrophages as well as other hematopoietic cells. Enzyme activity was increased fivefold and twofold in macrophages from BM and spleen, respectively, and could be precipitated with an antibody specific for human GC. Immunohistochemical analysis detected the human GC protein in 81% of the macrophages from five recipient mice. These data indicate that, after transduction of hematopoietic stem cells, the LG vector is capable of directing expression of human GC in the majority of macrophages from long-term reconstituted mice and producing enzyme levels comparable with endogenous mouse activity, suggesting that this virus may be useful in the treatment of Gaucher disease.

3T3 Cells↗
PubMed1992

Neonatal capsaicin-treatment in mice: effects on pancreatic peptidergic nerves and 2-deoxy-D-glucose-induced insulin and glucagon secretion.

It is not known whether sensory nerves are involved in the insulin, glucagon or glucose responses to autonomic nerve activation induced by 2-deoxy-D-glucose (2-DG). We therefore treated mice neonatally with capsaicin which permanently destroys sensory afferent nerve fibers. Immunohistochemistry of the pancreas at 13-14 weeks of age revealed a substantial reduction of calcitonin gene-related peptide (CGRP)-immunoreactive nerves and a partial reduction of substance P-immunoreactive nerves. In contrast, no effect was observed on galanin-immunoreactive nerves. At age 10-12 weeks, the mice were injected intravenously with 2-DG (500 mg/kg). In controls, 2-DG stimulated insulin and glucagon secretion and induced hyperglycemia (P less than 0.01). Capsaicin treatment partially reduced the glucose and glucagon responses to 2-DG (P less than 0.01). In contrast, the insulin response to 2-DG was not affected by capsaicin. It is concluded that the mouse pancreas contains capsaicin-sensitive sensory CGRP- and substance P-immunoreactive nerve fibers, whereas the galanin-immunoreactive nerve fibers are not sensitive to capsaicin. Furthermore, capsaicin-sensitive sensory nerve fibers are partially involved in 2-DG-induced glucagon secretion and hyperglycemia, whereas sensory nerves are not involved in 2-DG-induced insulin secretion.

Animals↗
PubMed1992

Effects of CCK-8 on the cytoplasmic free calcium concentration in isolated rat islet cells.

The C-terminal octapeptide of cholecystokinin (CCK-8) is known to stimulate insulin secretion. We examined its effects on the cytoplasmic free calcium concentration ([Ca2+]IC) in isolated rat pancreatic islet cells. At 8.3 mM glucose and 1.28 mM Ca2+, CCK-8 (100 nM) rapidly increased [Ca2+]IC to a short-lived peak, whereafter the [Ca2+]IC, within 1.5 minutes, fell to values below baseline. CCK-8 also rapidly increased the [Ca2+]IC at 3.3 mM glucose and in a calcium deficient medium. However, either at low glucose or in the absence of extracellular Ca2+, the post-peak [Ca2+]IC did not fall below baseline levels. The CCKA receptor antagonist, L-364,718 (20 nM), inhibited the effects of CCK-8 on [Ca2+]IC. The results suggest that CCK-8 in islet cells liberates calcium from intracellular stores by activating CCKA receptors.

Animals↗
PubMed1992

CCK-8-stimulated insulin secretion in vivo is mediated by CCKA receptors.

The effects of the cholecystokinin A (CCKA) receptor antagonist, L-364,718, and the CCKB receptor antagonist, L-365,260, on CCK-8-stimulated insulin secretion were studied in vivo in the mouse. It was found that CCK-8-stimulated insulin secretion was suppressed by L-364,718 at a low dose level (0.078 mumol/kg). In contrast, L-365,260 caused a partial inhibition of CCK-8-stimulated insulin release only at the high dose level (24 mumol/kg). It is concluded that the CCK-8-stimulated insulin release in vivo is mediated by CCK receptors of the CCKA subtype.

Animals↗
PubMed1992

Effects of intense chewing on some parameters of masticatory function.

This study recorded the influence of 30 minutes of intense chewing on the masticatory function in 17 healthy subjects. Functional recordings of mandibular movements and velocity, measurements of masticatory efficiency, maximal occlusal force, and occlusal force endurance time were performed before and after functional stimulation and finally in a 2- to 5-day follow-up registration. After intense chewing, a substantially decreased masticatory cycle and occlusal level phase duration were recorded. No major mandibular displacements or changes in velocity were recorded. Masticatory efficiency and maximal occlusal force in the incisal region diminished after 30 minutes of intense chewing. Occlusal force endurance time was also lowered after intense chewing. The measurements were close to their original values at the follow-up. Intense chewing had various short-term effects on masticatory function, including a significant reduction of masticatory efficiency, maximal occlusal force, occlusal force endurance, and masticatory cycle duration.

Adult↗