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Göran Andersson

Publications and source records attributed to Göran Andersson.

18 recordsLinked to original sources
PubMed2006

Cholesterol-sensing receptors, liver X receptor alpha and beta, have novel and distinct roles in osteoclast differentiation and activation.

UNLABELLED: The liver X receptor (alpha,beta) is responsible for regulating cholesterol homeostasis in cells. However, our studies using the LXRalpha-/-, LXRbeta-/-, and LXRalpha-/-beta-/- mice show that both LXRalpha and beta are also important for bone turnover, mainly by regulating osteoclast differentiation/activity. INTRODUCTION: The liver X receptors (alpha,beta) are primarily responsible for regulating cholesterol homeostasis within cells and the whole body. However, as recent studies show that the role for this receptor is expanding, we studied whether the LXRs could be implicated in bone homeostasis and development. MATERIALS AND METHODS: pQCT was performed on both male and female LXRalpha-/-, LXRbeta-/-, LXRalpha-/-beta-/-, and WT mice at 4 months and 1 year of age. Four-month-old female mice were additionally analyzed with reference to qPCR, immunohistochemistry, histomorphometry, transmission electron microscopy, and serum bone turnover markers. RESULTS: At the mRNA level, LXRbeta was more highly expressed than LXRalpha in both whole long bones and differentiating osteoblast-like MC3T3-E1 and osteoclast-like RAW 264.7 cells. Four-month-old female LXRalpha-/- mice had a significant increase in BMD because of an increase in all cortical parameters. No difference was seen regarding trabecular BMD. Quantitative histomorphometry showed that these mice had significantly more endosteal osteoclasts in the cortical bone; however, these cells appeared less active than normal cells as suggested by a significant reduction in serum levels of cross-linked carboxyterminal telopeptides of type I collagen (CTX) and a reduction in bone TRACP activity. Conversely, the female LXRbeta-/- mice exhibited no change in BMD, presumably because a significant decline in the number of the trabecular osteoclasts was compensated for by an increase in the expression of the osteoclast markers cathepsin K and TRACP. These mice also had a significant decrease in serum CTX, suggesting decreased bone resorption; however, in addition presented with an increase in the expression of osteoblast associated genes, bone formation markers, and serum leptin levels. CONCLUSIONS: Our findings show that both LXRs influence cellular function within the bone, with LXRalpha having an impact on osteoclast activity, primarily in cortical bone, whereas LXRbeta modulates trabecular bone turnover.

Acid Phosphatase↗
PubMed2005

Generation of therapeutic antibody responses against IgE in dogs, an animal species with exceptionally high plasma IgE levels.

Allergic diseases are common in dogs and the involvement of IgE in the pathogenesis of canine atopic dermatitis (CAD) and flea allergy dermatitis (FAD) is documented. However, many dogs do not achieve sufficient control of their allergic disease, and there is a great need for new treatment strategies. In order to address this issue we first needed to obtain a better picture of IgE-levels in dogs and how plasma IgE-levels are affected by breed, age and health status. IgE is normally present at diminutive concentrations in sera and detection by diagnostic methods has been a technical challenge. Here, we present a new in vitro assay for determining absolute levels of total IgE in sera from dogs. In a panel of 76 adult dogs diagnosed either as healthy or suffering from atopic dermatitis, autoimmunity or infection by skin parasites, we show that levels of IgE range from 1 to 41 microg/ml. This is almost 100 times the level observed in non-atopic humans. However, these exceptionally high IgE-levels in the dogs could not be correlated to either breed or health status. To address the issue of novel treatment strategies, the possibility of reducing the IgE-levels in nine Beagle dogs by immunization with a new therapeutic vaccine was investigated. High levels of anti-IgE antibodies were induced in all dogs, and the IgE-levels were subsequently decreased by a mean of 65%. This shows that the allergy vaccine is potent enough to break the tolerance against IgE, even when the initial IgE-levels are as high as those observed in dogs. Thus, the vaccination treatment may have the potential to serve as a future therapy for dogs with atopic diseases.

Age Factors↗
PubMed2005

Proteolytic excision of a repressive loop domain in tartrate-resistant acid phosphatase by cathepsin K in osteoclasts.

Tartrate-resistant acid phosphatase (TRAP) is a metallophosphoesterase participating in osteoclast-mediated bone turnover. Activation of TRAP is associated with the redox state of the di-iron metal center as well as with limited proteolytic cleavage in an exposed loop domain. The cysteine proteinases cathepsin B, L, K, and S as well as the matrix metalloproteinase-2, -9, -13, and -14 are expressed by osteoclasts and/or other bone cells and have been implicated in the turnover of bone and cartilage. To identify proteases that could act as activators of TRAP in bone, we report here that cathepsins K and L, in contrast to the matrix metalloproteinases, efficiently cleaved and activated recombinant TRAP in vitro. Activation of TRAP by cathepsin K/L was because of increases in catalytic activity, substrate affinity, and sensitivity to reductants. Processing by cathepsin K occurred sequentially by an initial excision of the loop peptide Gly(143)-Gly(160) followed by the removal of a Val(161)-Ala(162) dipeptide at the N terminus of the C-terminal 16-kDa TRAP subunit. Cathepsin L initially released a shorter Gln(151)-Gly(160) peptide and completed processing at Ser(145) or Gly(143) at the C terminus of the N-terminal 23-kDa TRAP subunit and at Arg(163) at the N terminus of the C-terminal 16-kDa TRAP subunit. Mutation of Ser(145) to Ala partly mimicked the effect of proteolysis on catalytic activity, identifying Ser(145) as well as Asp(146) (Funhoff, E. G., Ljusberg, J., Wang, Y., Andersson, G., and Averill, B. A. (2001) Biochemistry 40, 11614-11622) as repressive amino acids of the loop region to maintain the TRAP enzyme in a catalytically latent state. The C-terminal sequence of TRAP isolated from rat bone was consistent with cathepsin K-mediated processing in vivo. Moreover, cathepsin K, but not cathepsin L, co-localized with TRAP in osteoclast-resorptive compartments, supporting a role for cathepsin K in the extracellular processing of monomeric TRAP in the resorption lacuna.

Acid Phosphatase↗
PubMed2005

Phosphorylated osteopontin promotes migration of human choriocarcinoma cells via a p70 S6 kinase-dependent pathway.

This study examined the role of osteopontin (OPN), a phosphorylated secreted glycoprotein, in the promotion of trophoblastic cell migration, an early event in the embryo implantation process. Three human choriocarcinoma cell lines, namely JAR, BeWo, and JEG-3, were treated with variants of OPN differing in the extent of phosphorylation following sequential dephosphorylation with tartrate-resistant acid phosphatase (TRAP), and their migratory response was measured. The highly phosphorylated human milk form of OPN (OPN-1) strongly triggered migration in all three cell lines, whereas the less phosphorylated variants, OPN-2a and OPN-2b, failed to stimulate migration. JAR cell migration in response to OPN-1 was accompanied by a rapid rearrangement of actin filaments to the cellular membrane. Using broad spectrum protein kinase profiling, we identified p70 S6 kinase as a major signal transduction pathway activated by OPN-1 during the migratory response in JAR cells. Activation was blocked completely by rapamycin and LY294002, thus demonstrating that OPN-1-stimulated migration occurs through mTOR and PI3K pathways, respectively. Conversely, PD98059 did not affect the activation of p70 S6 kinase by OPN-1, therefore, this response does not involve the Ras/ MAPK signaling cascade. Together, these data show that the highly phosphorylated human OPN-1 can stimulate trophoblastic cell migration and provides evidence for the involvement of the PI3K/mTOR/p70 S6 kinase pathway in the JAR cells response. Because both OPN and TRAP are expressed in the uterus during early pregnancy, it is conceivable that extracellular phosphatases such as TRAP may modify OPN charge state and thus modulate cell migration.

Blotting, Western↗
PubMed2005

N-glycosylation influences the latency and catalytic properties of mammalian purple acid phosphatase.

Purple acid phosphatase (PAP), also known as tartrate-resistant acid phosphatase or uteroferrin, contains two potential consensus N-glycosylation sites at Asn(97) and Asn(128). In this study, endogenous rat bone PAP was found to possess similar N-glycan structures as rat recombinant PAP heterologously expressed in baculovirus-infected Sf9 insect cells. PAP from Sf9 cells was shown to contain two N-linked oligosaccharides, whereas PAP expressed by mammalian CHO-K1 cells was less extensively glycosylated. The extent of N-glycosylation affected the catalytic properties of the enzyme, as N97Q and N128Q mutants, containing a single oligosaccharide chain, exhibited a lower substrate affinity and catalytic activity compared to those of the fully glycosylated PAP in the native, monomeric state. The differences in substrate affinity and catalytic activity were abolished and partially restored, respectively, by proteolytic cleavage in the loop domain, indicating that the extent of N-glycosylation influences the interaction of the repressive loop domain with catalytically important residues.

Acid Phosphatase↗
PubMed2005

Polarization and secretion of cathepsin K precede tartrate-resistant acid phosphatase secretion to the ruffled border area during the activation of matrix-resorbing clasts.

The activation sequence of clasts (the designation clast was used because ultrastructurally in this tissue, it is not always possible to differentiate between chondroclasts sitting on cartilage and osteoclasts sitting on bone matrix) was studied in vivo using the healing of low-phosphate, vitamin D-deficiency rickets as a model system. Thus, the bones of 7-week-old rachitic animals were analyzed with a combination of morphological, biochemical, and molecular biological methods at 48 and 72 h, respectively, after change to normal food. A quantitative ultrastructural analysis showed that the number of clast profiles exhibiting the characteristic polarized features of actively resorbing cells, i.e., ruffled borders and clear zones, had reached normal levels after 48 h. By combining the data with quantitative analyses by the immunogold technique, we demonstrated that cathepsin K secretion was coupled to ruffled border formation in clasts irrespective of whether the number of polarized clasts was low (in rickets) or high (in healing). In contrast, the levels of tartrate-resistant acid phosphatase (TRAP) both between ruffles and in the outside matrix adjoining the ruffled border were low in polarized clasts both in rickets and at the early (48 h) healing time-point, but were increased at the latest (72 h) healing time-point. Interestingly, expression of TRAP and the cathepsin K at the mRNA level, as well as protein expression and the activity of TRAP, were not different during the healing sequence. Although the two enzymes are confined to the same clast populations, their secretion during the resorption process is apparently differentially regulated: cathepsin K secretion is coupled to ruffled border formation in clasts, whereas TRAP is secreted at a later stage during the resorption sequence, suggesting a role for secreted TRAP as a modulator of resorptive activity.

Acid Phosphatase↗
PubMed2005

Pig islet xenotransplantation: activation of porcine endogenous retrovirus in the immediate post-transplantation period.

BACKGROUND: Porcine endogenous retroviruses (PERV) are considered as the main infectious barrier in islet xenotransplantation. PERV has been shown to infect, but not to cause symptomatic disease in mice after islet transplantation. In vivo activation of PERV have so far not been examined. Expression of PERV was examined in adult and fetal porcine islets with or without the presence of known retroviral inducers or after transplantation to rats. METHODS: Isolated adult and fetal porcine islets were cultured under normal conditions or in the presence of dexamethasone or 5-azacytidine and 5-iodo-2-deoxyuridine. PERV mRNA content was analyzed by real-time quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and culture supernatants were analyzed for the presence of retroviral RT. Also, fetal islets were transplanted under the kidney capsule of immunocompetent or nude athymic rats. Expression of PERV mRNA in the grafts was evaluated by real-time quantitative RT-PCR. Infiltration of immunocompetent cells were evaluated by immunohistochemistry. RESULTS: Both fetal and adult islets in culture produced small or even undetectable amounts of PERV mRNA and retroviral RT. PERV expression was not enhanced by retroviral inducers. In contrast, activation of PERV expression was observed the first day after transplantation of fetal islet-like cell clusters in both athymic and normal rats. PERV expression peaked after 1 to 3 days and was then rapidly returned to background levels. PERV expression neither correlated with the innate immune response seen in athymic rats nor with the specific process of rejection in normal rats. CONCLUSION: Both fetal and adult islets produce low amounts of PERV mRNA in culture. After transplantation PERV expression is induced, seemingly independent of both the unspecific inflammatory response and the specific T-cell-mediated rejection process. It is speculated that PERV expression is correlated with the level of hypoxia in the islet xenograft.

Animals↗
PubMed2004

Intracellular machinery for matrix degradation in bone-resorbing osteoclasts.

UNLABELLED: In osteoclasts, TRACP co-localized with cathepsin K in transcytotic vesicles and was activated by cathepsin K in vitro, suggesting that TRACP may degrade organic matrix components in transcytotic vesicles in an event regulated by cathepsin K. INTRODUCTION: TRACP is an enzyme with unknown biological function. In addition to its phosphatase activity, TRACP is capable of generating reactive oxygen species (ROS). Bone-resorbing osteoclasts contain large amounts of TRACP, and transgenic animal models suggest that TRACP has a role in bone resorption. Osteoclasts resorb bone by secreting acid and lysosomal enzymes such as cathepsin K into an extracellular resorption lacuna between the cell membrane and bone surface. Matrix degradation products are then endocytosed, transcytosed, and secreted through a functional secretory domain in the basolateral membrane facing bone marrow. MATERIALS AND METHODS: We have studied intracellular localization of TRACP in osteoclasts with antibodies against various known endosomal and lysosomal proteins using confocal microscopy. We also studied co-localization of TRACP with cathepsin K and endocytosed bone matrix components and the effect of cathepsin K digestion on the ROS generating activity of TRACP in vitro. RESULTS: Double-staining experiments of TRACP with endosomal and lysosomal markers showed that, although some endosomal staining was detected, TRACP was not present in lysosomes. However, TRACP was present in transcytotic vesicles, where it co-localized with cathepsin K. Cathepsin K digestion of TRACP in vitro increased the phosphatase activity by 5.6-fold and the ROS generating activity by 2.0-fold. CONCLUSIONS: These results suggest that cathepsin K may activate the ROS-generating activity of TRACP in transcytotic vesicles of resorbing osteoclasts, the ROS being targeted to finalize degradation of organic bone matrix components during their transcytosis.

Acid Phosphatase↗
PubMed2004

IGF2 antisense transcript expression in porcine postnatal muscle is affected by a quantitative trait nucleotide in intron 3.

A paternally expressed quantitative trait locus (QTL) for muscle mass was mapped to the insulin-like growth factor II (IGF2) locus at the distal end of pig chromosome 2p. It was recently demonstrated that a G to A transition at position intron 3-nt 3072 is the quantitative trait nucleotide (QTN) underlying this QTL. In this study we report for the first time the existence of imprinted porcine IGF2 antisense transcripts (IGF2-AS) and demonstrate that their expression in postnatal muscle is also affected by the QTN. A coregulated expression of IGF2 and IGF2-AS RNAs in muscle and liver with decreasing transcription from fetal to adult age was demonstrated. Further, the significant difference found in expression of IGF2-AS in postnatal muscle between individuals with different QTL genotypes and the lack of significant differences in fetal muscle and liver reflect completely what has been found for IGF2 sense transcripts.

Animals↗
PubMed2003

A regulatory mutation in IGF2 causes a major QTL effect on muscle growth in the pig.

Most traits and disorders have a multifactorial background indicating that they are controlled by environmental factors as well as an unknown number of quantitative trait loci (QTLs). The identification of mutations underlying QTLs is a challenge because each locus explains only a fraction of the phenotypic variation. A paternally expressed QTL affecting muscle growth, fat deposition and size of the heart in pigs maps to the IGF2 (insulin-like growth factor 2) region. Here we show that this QTL is caused by a nucleotide substitution in intron 3 of IGF2. The mutation occurs in an evolutionarily conserved CpG island that is hypomethylated in skeletal muscle. The mutation abrogates in vitro interaction with a nuclear factor, probably a repressor, and pigs inheriting the mutation from their sire have a threefold increase in IGF2 messenger RNA expression in postnatal muscle. Our study establishes a causal relationship between a single-base-pair substitution in a non-coding region and a QTL effect. The result supports the long-held view that regulatory mutations are important for controlling phenotypic variation.

Animals↗
PubMed2003

Differential expression of osteopontin and bone sialoprotein in bone metastasis of breast and prostate carcinoma.

Breast and prostate cancer often metastasise to the skeleton. Interestingly, the histopathological characteristics of the bone lesions that arise from these two cancer types differ. Breast tumours give rise to metastases in the skeleton with a mixed lytic/sclerotic pattern, whereas a predominantly sclerotic pattern is seen in metastases from prostate tumours. Osteopontin (OPN) and bone sialoprotein (BSP) are bone matrix proteins that have been implicated in the selective affinity of cancer cells for bone. In the present study, 21 patient cases with skeletal metastasis and their respective primary tumours (12 with breast cancer, 9 with prostate cancer) were investigated by immunohistochemistry in order to assess the level of OPN and BSP. Moderate to strong OPN expression was found in 42% of all breast tumours and in 56% of all prostate tumours. Significantly more breast cancer bone metastases exhibited high OPN expression, 83%, as compared with prostate tumour bone metastases, 11% (P = 0.0019). In contrast, moderate to strong BSP expression was found in 33% of breast tumours and in 89% of prostate tumours. In the bone lesions, only 33% of breast tumour metastases showed moderate/strong BSP expression compared to 100% of prostate tumour metastases (P = 0.0046). This divergent pattern of OPN/BSP expression could be an important determinant for the different characteristics of these two types of bone metastasis, i.e., lytic vs. sclerotic, consistent with the proposed role of OPN in differentiation and activation of osteoclasts and of BSP as a stimulator of bone mineralisation.

Adult↗
PubMed2003

Doxorubicin-resistant, MRP1-expressing U-1285 cells are sensitive to idarubicin.

A doxorubicin-resistant subline (U-1285dox(900)) was derived from the human small cell lung carcinoma cell line U-1285. U-1285dox(900) was exposed to a wide range of anticancer agents to determine its resistance profile. In contrast to U-1285 cells, the resistant subline U-1285dox(900) expressed elevated MRP1 mRNA detected by reversed transcriptase-polymerase chain reaction (RT-PCR) and MRP1 protein analyzed with Western blot. Neither MDR1 mRNA nor P-glycoprotein could be detected in the parental cell line or resistant subline. U-1285dox(900) exhibited high resistance to doxorubicin, epirubicin, daunorubicin, and vincristine, an intermediate resistance to mitoxantrone, and a low resistance to etoposide. A collateral sensitivity to cytosine arabinoside, chlorodeoxyadenosine, and melphalan was observed. The resistance could be reversed by buthionine-sulphoximine and verapamil for all tested drugs. Compared with daunorubicin, resistance to idarubicin was very low, 14-fold and 2.6-fold, respectively. This was associated with a higher accumulation due to a slower transport of idarubicin out of U-1285dox(900) cells.

Area Under Curve↗
PubMed2003

TRACP as an osteopontin phosphatase.

TRACP is synthesized as a latent proenzyme requiring proteolytic processing to attain maximal phosphatase activity. Excision of an exposed loop domain abolishes the interaction between the loop residue Asp146 and a ligand to the redox-sensitive iron of the active site, most likely Asn91, providing a mechanism for the enzyme repression. Both cathepsin K and L efficiently cleave in the loop domain and activate the latent enzyme, and we propose that cathepsin K acts as a physiological activator of TRACP in osteoclasts, whereas cathepsin L might fulfill a similar role in different types of macrophages. Considering the rather broad substrate specificity of TRACP, a tight regulation of its activity in the cell appears warranted. Besides proteolytic cleavage, the enzyme should need a specific local environment with a slightly acidic pH and reducing equivalents to keep the enzyme fully active. Cellular subcompartments where these required conditions prevail are potential subcellular site(s) of TRACP action. Of bone phosphoproteins shown to be substrates for TRACP, both osteopontin and bone sialoprotein are colocalized with TRACP in the resorption lacuna of the osteoclasts, and dephosphorylation of OPN impair its ability to promote adhesion as well as migration of osteoclasts in vitro. A role for TRACP as an osteopontin phosphatase in bone is therefore suggested. The expression of TRACP as well as OPN in other tissues with possible interactions between the two could suggest a more general function for TRACP as a regulator of OPN phosphorylation and bioactivity.

Acid Phosphatase↗
PubMed2003

The effects of colony-stimulating factor-1 (CSF-1) on the development of osteoclasts and their expression of tartrate-resistant acid phosphatase (TRAP) in toothless (tl-osteopetrotic) rats.

The osteopetrotic mutation toothless (tl) in the rat is characterized by a limited number of osteoclasts with reduced amounts and/or activity of tartrate-resistant acid phosphatase (TRAP). Treatment of tl/tl mutants with the cytokine colony-stimulating factor (CSF)-1 increases both osteoclast number and enzyme activity, consistent with a loss-of-function mutation in the CSF-1 gene recently detected in this mutant. We have pursued these observations to demonstrate that there is a dose-dependent increase in osteoclast number, but not to normal levels. Osteoclasts in CSF-1-treated tl/tl mutants are large, have well-developed clear zones and ruffled borders, and secrete TRAP into resorption lacunae. The expression of TRAP mRNA, protein, and enzyme activity per bone appear normal after CSF-1 treatment. However, in contrast to the predominantly apical intracellular distribution in normal osteoclasts, an enrichment of TRAP enzyme activity in osteoclasts of CSF-1-treated tl/tl mutants is observed in the basal part of the cell. Our observations suggest that the CSF-1-treated mutant bones contain an abundance of mature osteoclasts, actively expressing markers for osteoclasts such as TRAP, cathepsin K, and matrix metalloproteinase (MMP)-9. Accumulation of TRAP at the end of the endocytic pathway in mature osteoclasts formed during CSF-1 treatment suggests that the TRAP enzyme has a rapid turnover in these highly active cells and uses a transcytotic pathway.

Acid Phosphatase↗
PubMed2002

Osteoclasts from mice deficient in tartrate-resistant acid phosphatase have altered ruffled borders and disturbed intracellular vesicular transport.

Tartrate-resistant acid phosphatase (TRAP) is an enzyme highly expressed in osteoclasts (OC) and chondroclasts. As an approach to pinpoint the function of TRAP in bone-resorbing osteoclasts, the morphological phenotypic alterations of bone and osteoclasts in mice with targeted disruption of the TRAP gene were assessed by quantitative histomorphometry and immunocytochemistry at the light microscopic and ultrastructural levels. TRAP-deficient mice display alterations in the epiphyseal growth plates as evidenced by increased height with disorganized columns of chondrocytes, in particular affecting the zone of hypertrophic chondrocytes, consistent with a disturbance of chondrocyte maturation and chondroclastic resorption at the epiphyseal/metaphyseal junction. TRAP -/- mice express an early onset osteopetrotic bone phenotype, apparent already at 4 weeks of age. The differentiation of OCs was apparently normal; however, the osteoclasts in TRAP-deficient mice were less active in terms of degradation or release of the resorption marker C-terminal type I collagen cross-linked peptide, indicative of an intrinsic defect. Ultrastructural morphometry disclosed that OCs from TRAP-deficient young mice exhibited an increased relative area of ruffled borders. Moreover, mutant OC accumulated cytoplasmic vesicles 200-500 nm in size in both ruffled border and basolateral parts of the cytoplasm, reflecting disturbed intracellular transport. The accumulated vesicles were not likely derived from the secretory pathway, since cathepsin K was detected at normal levels in the ruffled border area and matrix in TRAP -/- mice. In summary, the resorptive defect in TRAP-deficient OCs is reflected by a disturbance at the level of ruffled borders and intracellular transport vesicles. Consequently, accumulation of vesicles in the cytoplasm of mutant OCs indicates a novel function for TRAP in modulating intracellular vesicular transport in osteoclasts.

Acid Phosphatase↗
PubMed2002

Elucidation of estrogen receptor function in bone with the use of mouse models.

Since the discovery that estrogen receptors (ERs) are present in bone cells, there has been intense research into the action of estrogen in bone. During the past decade, humans with disturbed estrogen signaling, either as a result of ER alpha or aromatase deficiency, have been reported. Furthermore, mouse models have been established with a deficiency of ER alpha, ER beta or both, in addition to deficiency of aromatase. This review focuses on data accumulated during the past three years from studies of knockout mice with impaired estrogen signaling resulting from ER or aromatase deficiency.

Animals↗
PubMed2002

Expression of xenogeneic MHC class II molecules in HLA-DR(+) and -DR(-) cells: influence of retrovirus vector design and cellular context.

We recently established that molecular chimeras of major histocompatibility complex (MHC) class II molecules, created via retroviral transfer of allogeneic class II cDNAs into bone marrow cells (BMCs), alleviated complications associated with mixed BMC chimeras while leading to T cell tolerance to renal grafts sharing the transferred class II. Initially demonstrated for allogeneic transplants in miniature swine, this concept was extended to T-dependent antibody (Ab) responses to xenogeneic antigens (Ags) in the pig --> baboon combination. Successful down-regulation of T cell responses appeared, however, to be contingent on a tight lineage-specific expression of transferred class II molecules. The present studies were, therefore, designed to evaluate the influence of construct design and cellular environment on expression of retrovirally transferred xenogeneic class II cDNAs. Proviral genomes for pig class II SLA-DR expression, differing only at the marker neo(r) or enhanced green fluorescent protein (EGFP) gene, showed increased membrane SLA-DR density on HLA-DR(-) fibroblasts as well as HLA-DR(+), TF-1 erythroleukemia cells. More importantly, HLA-DR(+) human B cell lines, although efficiently transduced with pig DR retroviruses, exhibited unstable surface pig DR. Surface pig DR- B cells, nevertheless, stimulated autologous human T cells pre-sensitized to pig Ags, a proliferation likely occurring through presentation of class II-derived peptides. Collectively, these data suggest that surface expression of transferred class II molecules is not related to the ability of recipient cells to synthesize xenogeneic class II molecules but rather to their Ag processing capacities.

Animals↗