| 11. |
- Georgii-Hemming, Patrik
(författare)
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Life, death ant the role of IGF-I in human multiple myeloma
- 1998
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Multiple myeloma (MM) is a clonal expansion of malignant cells with a plasmablast- plasma cell morphology in the bone marrow. It is a fatal disease with a median survival of 2-3 years after start of conventional therapy. The aim of the thesis was to study the regulation of growth and survival of MM cell lines and primary, cells to identify potential targets for therapy. The results of the thesis show that MM cells express IGF-I (insulin-like growth factor-I) receptors and IGF-I. Furthermore, IGF-I was shown to stimulate growth and survival of MM cells. When IGF-IR signaling is inhibited by anti-IGF-IR antibodies in MM cells they are growth inhibited and may also undergo apoptosis. The somatostatin analogue octreotide has been demonstrated to interfere with the action of IGF-I. The results of the thesis show that MM cells express sst2, sst3 and sst5 which bind octreotide with a moderate or high affinity. Moreover, octreotide inhibits the growth of all investigated MM clones. In a few cases growth inhibition was also accompanied by the induction of apoptosis.Resistance to apoptosis may be important for cell survival and drug resistance in MM clones. Studies in the thesis demonstrated that interferon (IFN)-α and IFN-γ augment the sensitivity to Fas-induced apoptosis in MM cells independently of their growth inhibitory effect. The sensitivity to apoptosis was also increased by inhibition of IGF-IR signaling. Incubation of MM cells with anti-IGF-IR antibodies increased the sensitivity of the cells to apoptosis induced by Fas ligation and the glucocorticoid dexamethasone. It can be concluded that the IGF-IR may be a potential target for therapy in MM. Furthermore, increasing the sensitivity of MM cells to apoptosis by treatment with IFNs or drugs that interfere with the action of IGF-I may increase the sensitivity of the tumor cells to cytotoxic drugs.
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| 12. |
- Karlsson, Mikael
(författare)
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Fc receptors and feedback regulation by antibodies
- 1999
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The purpose of this investigation was to determine the role of Fc receptors for IgG (FcγRI, FcγRIIB, FcγRIII and FcRn) in immunoregulation. IgG is known to enhance the response to soluble antigens such as bovine serum albumin whereas the response to particulate antigens is suppressed.The ability of IgG to induce specific suppression of immune responses has been used clinically to prevent haemolytic disease of the newborn (Rhesus prophylaxis). The favoured mechanism behind IgG-mediated suppression has been specific inhibition of antibody production through an inhibitory Fc receptor for IgG, FcγRIIB. We here study the importance of Fc receptors for IgG using Fc receptor-deficient mice immunised with sheep red blood cells alone or together with IgG anti-sheep red blood cells. IgG was able to suppress the primary response in all Fc receptor-deficient mice measured as number of sheep red blood cell specific B cells. In addition, IgE and F(ab')2 fragments of IgG induced antigen-specific suppression, suggesting an Fc-independent mechanism involving masking of epitopes. FcγRIIB was required for suppression of secondary antibody responses in vitro but not in vivo.To study the enhancing effect by IgG, the Fc receptor-deficient mice were immunised with haptenated bovine serum albumin alone or together with hapten-specific IgG. The resulting enhancement was measured as specific bovine serum albumin antibody in serum. We found that in order for IgG to enhance the response to soluble antigen there is a requirement for the common-y chain which is a part of FcγRI and FcγRIII. Using mice specifically deficient for FcγRIII we find that IgG can enhance the response pointing towards FcγRI as the receptor responsible for the effect. An additional finding was that theenhancement was greatly elevated in mice lacking FcγRIIB. This suggests that FcγRIIB has a function in preventing antibody responses from exceeding a certain level rather than in causing complete suppression.
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| 13. |
- Spets, Helena
(författare)
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Regulation of apoptosis in human multiple myeloma
- 1998
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Multiple myeloma (MM) is a clonal expansion of malignant plasmablasts-plasma cells in the bone marrow. MM is slowly proliferating and an increased resistance to apoptosis may be crucial for the tumor progression. By the use of MM cell lines and primary cells the role of some growth factors, and genes regulating survival or apoptosis in human MM cell lines and primary cells, was investigated. The in vitro requirements for growth and survival of MM cells are heterogenous. This has hampered studies of molecular mechanisms underlying the effects of interleukin-6 (IL-6). The results of this thesis showthat IL-6, in the presence of serum, can convey survival dissociated from the effects of IL-6 on proliferation in two pairs of MM cell lines, U-266-1970/U-266-1984 and HL407E/HL407L. In the absence of serum IL-6 may also augment apoptosis. The findings therefore provide a useful human model system of IL-6 dependent and independent cell lines to study the molecular mechanisms underlying IL-6 effects in MM.The thesis also report on the expression of seven Bcl-2 related proteins; Bcl-2, Mcl-1, Bcl-x, Bcl-w, Bax, Bak, and Bad in MM cell lines and normal plasma cells. Elevated expression levels of Bcl-2 and Mel-1, as well as decreased levels of Bax expression were demonstrated in MM cells, and may therefore be associated with the malignant phenotype. Downregulation of the expression of Bcl-2, Mcl-1, and Bcl-x, and an upregulated Bax expression were recorded as a result of IL-6 deprivation in two IL-6 dependent-MM cell lines.A poor response to Fas and dexamethasone induced apoptosis is evident in some MM cell lines and primary cells. We here report that sensitivity to Fas mediated apoptosis can be augmented by IFN-γ or IFN-α in the U-266-1970, U-266-1984, and U-1958 cell lines. The mechanism by which IFNs regulate the sensitivity to apoptosis is unknown. IFNs increased the Fas antigen expression in one of three cell lines, but did not alter the expression of Bcl-2, or Bax, and was shown to be independent of growth inhibition.We also present results showing that IGF-IR signaling can mediate resistance to anti-Fas and dexamethasone induced apoptosis in two IGF-I responsive MM cell lines, LP-1 and Karpas 707. The results show that IGF-IR signaling may be a potent inhibitor of apoptosis, but does not counteract the growth inhibitory effect of dexamethasone in MM cell lines. Taken together this thesis provides important insights in the regulation of cell survival in MM cells by IL-6 and IGF-I, and suggests possible regulators of sensitivity to induced apoptosis in MM.
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| 14. |
- Thunberg, Ulf
(författare)
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Detection of immunoglobulin heavy chain gene rearrangement with PCR for MRD analysis in lymphoproliferative disorders
- 1998
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Immunoglobulin heavy chain (IGH) gene rearrangement occurs during early B-lymphocyte differentiation, assembling the different IGH gene segments to a functional gene, which can serve as a marker for study of lineage association and detection of Minimal Residual Disease (MRD) in clonal diseases deriving from B-lymphocytes or their early differentiation stages. Use of a molecular marker for the leukemic cells could help improve treatment by monitoring therapeutic efficacy, predicting relapse, and identifying very small amounts of tumour cells contaminating autografts after purging or enrichment of stem cells.This work focused on the task of improving the specificity of clonality and MRD tests by developing simple laboratory methods based on Polymerase Chain Reaction (PCR) with which to distinguish monoclonal from polyclonal cells. These methods were compared with conventional histopathological and immunocytological techniques in a retrospective study concerning the possibility of predicting recurrence of childhood pre-B Acute Lymphoblastic Leukemia (ALL). Clonal evolution in childhood pre-B ALL was also studied by comparing the rearrangements at diagnosis versus at relapse. A highly sensitive clonospecific PCR-method was utilized to detect small amounts of monoclonal cells before and after stem cell purging in mobilized peripheral blood stem cells from multiple myeloma patients.
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| 15. |
- Uhrbom, Lene
(författare)
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Roles of p16INk a and PDGF-B in glioma
- 1999
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Gliomas are diffuse astrocytic tumors of the central nervous system (CNS) which usually affect adults. This thesis work has focused on molecular mechanisms of altered growth control in glioma, and the significance of two gene products belonging to pathways commonly affected in glioma, p16INK4a and PDGF-B, has been investigated in vitro and in vivo.The INK4a locus is one of the most frequently lossed tumor suppressor gene of human gliomas. It encodes a G1 specific cell cycle inhibitor, p16INK4a, which was ectopically expressed in the human malignant glioma cell line U-1242 MG. Forced expression caused a G1 arrest and senescence in the cells, defined by an inability to induce DNA replication at subconfluency in the presence of serum, an enlarged cell morphology and expression of a senescence marker, senescence-associated β-galactosidase (SA-β-gal).Many studies have implicated a role for platelet-derived growth factor (PDGF) in human glioma. We established a mouse brain tumor model by injecting a murine retrovirus containing the PDGP-B cDNA together with wild-type virus, intracranially in neonatal mice. The induced malignant brain tumors displayed features of human gliomas. Tumor cells were nestin positive and expressed PDGP-B and PDGF-Rα mRNA, suggesting an autocrine stimulation of a neuroepithelial progenitor cell as the mechanism of tumor induction. A cell line was established from one PDGF-B induced tumor. Cultured tumor cells expressed PDGF-B causing autophosphorylation of endogenous PDGF receptors. A specific PDGF tyrosine kinase inhibitor caused a striking inhibition of PDGF receptor tyrosine phosphorylation and cell proliferation, showing a dependence of PDGF stimulation in initiation as well as progression of these tumors. Furthermore, the contribution of insertional mutagenesis in tumor development was analysed. Retroviral insertions in genomic DNA from 15 tumors were investigated with inverse polymerase chain reaction (IPCR) and sequencing. Preliminary data indicate the presence of a common integration site in a sequence mapped to a defined region of human chromosome 22, corresponding to mouse chromosome 15.
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| 16. |
- Wester, Kenneth
(författare)
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Quantitative immunochemistry in tissue sections : With special reference to urinary bladder carcinoma
- 1999
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Urinary bladder carcinoma affects about 2 000 people in Sweden every year. It is a heterogeneous disease and for adequate treatment decisions, prognostic tools are needed. Many of these are based on immunohistochemistry (MC) and hampered by poorly reproducible methodologies and subjective evaluations.This investigation aimed to evaluate and define the conditions for an objective and reproducible quantification of(MC) applied to routinely processed (formalin-fixed and paraffin-embedded) tissue material. To establish theseconditions, proliferation-related proteins, tumour-suppressor proteins and endothelial cell markers were studiedThree computerised colour-based image analysis methods, aiming to reduce user interaction, were developed and evaluated for variability and reproducibilty in quantification of IHC applied to histological sections. Themethods proved stable to variations in focus- and light-settings but revealed a sensitivity to poor IHC-qualityand choice of chromogen. Implementation of an external control system, used as base for computing theclassifier, further increased the stability.Using these methods, effects of varying tissue-handling and immunostaining conditions were analysed. Storageof both paraffin-blocks and -sections was deleterious to immunoreactivity but could be compensated for bytayloring of the heat induced epitope retrieval (HIER) protocol.A control system for MC based on cultured cells was developed to reflect variations in tissue-handling and IHC-conditions. Control and biopsy material showed similar sensitivity to section storage, MC- and HIER-protocolvariations. Furthermore, controls reflected variations in fixation time and proved stable to variations in sectionthickness.
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| 17. |
- Zhang, Wei Wei
(författare)
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Microangiopathies of the human brain b immunohistochemical studies on extracellular matrix components in arterial vessels and endothelin
- 1997
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Microangiopathies may cause ischemic brain lesions and are of fundamental importance in vascular dementia. Risk factors include high age, hypertension, diabetes and Alzheimer's disease. In addition, recent studies have focused on autosomal dominant types of arteriopathy causing leukoencephalopathy,psychiatric disturbances, stroke and dementia (CADASIL). This thesis concerns various collagens andbasal lamina components which are deposited in vascular walls of cases presenting cerebral microangiopathy. In addition, endothelin-like immunoreactivity has been studied in CADASIL cases andsome other brain diseases.CADASIL cases described by Sourander and Wålinder (1977) were re-investigated. Those with longduration of the disease presented marked expression of fibrillary collagen types I, Ill, V and VI and of thebasal lamina components, collagen type IV and laminin. Deposits appeared also in non-familial casespresenting hyalinosis and in cases with the Binswanger type of leukoencephalopathy. Media degeneration and deposition of fibrillary collagens will in CADASIL, Binswanger's disease and hyalinosis graduallytransform the vessels to obliterated stiff tubings which can not properly regulate blood flow. On the otherhand, many vessels in cases with amyloid angiopathy did not show accumulations of collagens I, Ill and V. Such vessels may be more prone to rupture and cerebral hemorrhages than those in which fibrillarycollagens have been deposited. Finally, studies on CADASIL cases, other cases with brain infarcts, lacunas and Alzheimer's disease demonstrated an endothelin-like immunoreactivity in reactive perivascular astrocytes. The idea was therefore proposed that during the course of brain lesions activated astrocytes start a production of endothelin-1 which after release may reach nearby microvessels and inducevaseconstriction leading to additional brain injury.Structural as well as humoral factors are both most likely important for the production of brain lesionsin cases presenting cerebral microangiopathies.
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| 18. |
- van Dijk-Härd, Iris, et al.
(författare)
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Age-related impaired affinity maturation and differential D-JH gene usage in human VH6-expressing B lymphocytes from healthy individuals
- 1997
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Ingår i: European Journal of Immunology. - : Wiley-VCH Verlagsgesellschaft. - 0014-2980 .- 1521-4141. ; 27:6, s. 1381-1386
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Tidskriftsartikel (refereegranskat)abstract
- To elucidate the basic molecular events underlying humoral immunity during ontogeny and senescence, we analyzed a panel of 179 polymerase chain reaction-derived VH6-D-JH rearrangements from cord blood, peripheral blood, and spleen. Nucleotide sequence analysis of the CDR3 region shows that there is a difference in D and JH gene usage in functional rearrangements between lymphocytes from peripheral blood and spleen. Analysis of the VH6 gene shows that the mutational frequencies rise from 0.81% in cord blood to 1.96% in peripheral blood lymphocytes derived from young adults, and decrease to 0.80% in samples from individuals older than 50 years. The number of rearrangements carrying mutations follows a similar pattern: 22% in cord blood, 73% in the age group 20-49 years, and 57% in the age group over 50 years. The mutational frequencies among the mutated genes are, however, similar for cord blood and young adults, 2.76% and 2.51%, respectively, and 1.3% in older adults. These data show an age-related impaired affinity maturation which might relate to the decrease in immunological responsiveness among the elderly.
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| 19. |
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| 20. |
- Blomquist, H K, et al.
(författare)
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Glycerol kinase deficiency in two brothers with and without clinical manifestations.
- 1996
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Ingår i: Clinical genetics. - 0009-9163 .- 1399-0004. ; 50:5, s. 375-9
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Tidskriftsartikel (refereegranskat)abstract
- We report two brothers with glycerol kinase deficiency (GKD). The older brother had serious clinical symptoms, mental and growth retardation, abnormal skeleton, spontaneous fractures and premature loss of abnormal teeth. He and his mother had low serum phosphate levels. He had elevated serum and urine glycerol levels and GKD was found in cultured fibroblasts. Prenatal diagnosis was performed in the second pregnancy. Glycerol kinase activity was considered normal in a chorionic villus sample of the foetus. After birth, it was found that the boy had elevated serum and urine glycerol levels. Enzymatic analysis in cultured fibroblasts revealed that this boy also had GKD, in spite of having no expression of the disease. Chromosomal analyses in the parents and both boys were normal. Major rearrangements or deletions were not detected in molecular studies of DNA from the two brothers. The hybridisation pattern was normal and no allelic loss was observed.
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