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1.
J Inflamm Res ; 7: 121-31, 2014.
Article in English | MEDLINE | ID: mdl-25258549

ABSTRACT

B-cell activating factor (BAFF) is a B-cell survival factor with a key role in B-cell homeostasis and tolerance. Dysregulated BAFF expression may contribute to autoimmune diseases or B-cell malignancies via effects on abnormal B-lymphocyte activation, proliferation, survival, and immunoglobulin secretion. Monoclonal antibodies were generated against human BAFF, characterized for species specificity and affinity, and screened for the ability to neutralize both membrane-bound and soluble BAFF. In addition, studies were undertaken to determine the relative potency of membrane-bound and soluble BAFF. Tabalumab has a high affinity for human, cynomolgus monkey, and rabbit BAFF. No binding to mouse BAFF was detected. Tabalumab was able to neutralize soluble human, cynomolgus monkey, or rabbit BAFF with equal potency. Our data demonstrate that membrane-bound BAFF can be a more potent stimulus for B-cells than soluble BAFF, and tabalumab also neutralized membrane-bound BAFF. Tabalumab prevented BAFF from binding to BAFF receptors and demonstrated pharmacodynamic effects in human BAFF transgenic mice. Tabalumab is a high-affinity human antibody with neutralizing activity against membrane-bound and soluble BAFF. Given our findings that membrane-bound BAFF can have greater in vitro potency than soluble BAFF, neutralization of both forms of BAFF is likely to be important for optimal therapeutic effect.

2.
Transgenic Res ; 13(4): 385-96, 2004 Aug.
Article in English | MEDLINE | ID: mdl-15517997

ABSTRACT

It has been demonstrated that GSK3beta is involved in Alzheimer Disease (AD) pathogenesis. In order to understand the underlying mechanism, we have generated and characterized transgenic mice in which the constitutively active human GSK3beta (with S9A mutation) was overexpressed in the brain under the control of the platelet-derived growth factor (PDGF) B-chain promoter. Varying levels of human GSK3betaS9A transgene protein expression was observed in six of the seven founders generated. Line 3083, 3107, 3112 and 3125 displayed higher GSK3betaS9A protein expression levels. Immunostaining analysis demonstrated that transgene expression was observed mainly in cortex and hippocampus of transgenic brain. Expression of human GSK3beta transgene did not significantly change the brain total GSK3beta protein levels in any of the generated mouse lines, as comparing to age matched wild type mice. Although significant kinase activity was detected in human GSK3betaS9A transgene protein extracted from brains of all six expressing lines, significant increase in total GSK3betaS9A kinase activity was observed only in the offspring of line 3083 and 3107. By analyzing the offspring from several transgenic mouse lines, including lines other than 3083 and 3107, it was found that overexpressed constitutively active human GSK3betaS9A resulted in hyperphosphorylation of tau and morphology reminiscent of pretangle-like neurons in cortex and hippocampus.


Subject(s)
Brain/pathology , Glycogen Synthase Kinase 3/genetics , Neurons/pathology , tau Proteins/metabolism , Amino Acid Substitution , Animals , Becaplermin , Brain/enzymology , Cerebral Cortex/enzymology , Cerebral Cortex/pathology , Disease Models, Animal , Glycogen Synthase Kinase 3/chemistry , Glycogen Synthase Kinase 3 beta , Hippocampus/enzymology , Hippocampus/pathology , Humans , Mice , Mice, Transgenic , Neurofibrillary Tangles/pathology , Neurons/enzymology , Phosphorylation , Platelet-Derived Growth Factor/genetics , Promoter Regions, Genetic/genetics , Proto-Oncogene Proteins c-sis , tau Proteins/genetics
3.
Biochemistry ; 43(22): 6899-908, 2004 Jun 08.
Article in English | MEDLINE | ID: mdl-15170327

ABSTRACT

Lithium is one of the most widely used mood-stabilizing agents for the treatment of bipolar disorder. Although the underlying mechanism(s) of this mood stabilizer remains controversial, recent evidence linking lithium to neurotrophic/neuroprotective effects (Choi and Sung (2000) 1475, 225-230; Davies et al. (2000) 351, 95-105) suggests novel benefits of this drug in addition to mood stabilization. Here, we report that both lithium as well as valproic acid (VPA) inhibit beta-amyloid peptide (Abeta) production in HEK293 cells stably transfected with Swedish amyloid precursor protein (APP)(751) and in the brains of the PDAPP (APP(V717F)) Alzheimer's disease transgenic mouse model at clinically relevant plasma concentrations. Both lithium and VPA are known to be glycogen synthase kinase-3 (GSK3) inhibitors. Our studies reveal that GSK3beta is a potential downstream kinase, which modulates APP processing because inhibition of GSK3 activity by either a dominant negative GSK3beta kinase-deficient construct or GSK3beta antisense oligonucleotide mimics lithium and VPA effects. Moreover, lithium treatment abolished GSK3beta-mediated Abeta increase in the brains of GSK3beta transgenics and reduced plaque burden in the brains of the PDAPP (APP(V717F)) transgenic mice.


Subject(s)
Alzheimer Disease/metabolism , Amyloid beta-Protein Precursor/metabolism , Lithium/pharmacology , Protein Processing, Post-Translational , Alzheimer Disease/drug therapy , Animals , Bipolar Disorder/drug therapy , Brain/metabolism , Cells, Cultured , Enzyme Inhibitors/pharmacology , Genes, Dominant , Glycogen Synthase Kinase 3/antagonists & inhibitors , Glycogen Synthase Kinase 3/genetics , Glycogen Synthase Kinase 3/metabolism , Glycogen Synthase Kinase 3 beta , Humans , Mice , Mice, Transgenic , Oligonucleotides, Antisense/pharmacology , Valproic Acid/pharmacology
4.
Blood ; 102(9): 3206-9, 2003 Nov 01.
Article in English | MEDLINE | ID: mdl-12855566

ABSTRACT

In a search for novel growth factors, we discovered that human interleukin-20 (IL-20) enhanced colony formation by CD34+ multipotential progenitors. IL-20 had no effect on erythroid, granulocyte-macrophage, or megakaryocyte progenitors. IL-20 transgenic mice increased the numbers and cell cycling of multipotential but not other progenitors. IL-20 administration to normal mice significantly increased only multipotential progenitor cells, demonstrating that IL-20 significantly influences hematopoiesis, with specificity toward multipotential progenitors. This is the first cytokine with such specificity identified.


Subject(s)
Hematopoietic Stem Cells/cytology , Interleukins/pharmacology , Multipotent Stem Cells/drug effects , Animals , Antigens, CD34 , Bone Marrow Cells/cytology , Cell Division/drug effects , Cells, Cultured , Fetal Blood/cytology , Hematopoiesis/drug effects , Hematopoietic Stem Cells/drug effects , Humans , Interleukins/administration & dosage , Mice , Mice, Transgenic , Multipotent Stem Cells/cytology
5.
Int Immunol ; 15(7): 861-70, 2003 Jul.
Article in English | MEDLINE | ID: mdl-12807825

ABSTRACT

LIGHT, a newly identified member of the tumor necrosis factor (TNF) family, is expressed on activated T lymphocytes. To evaluate how LIGHT contributes to T cell functions, we generated LIGHT-deficient (LIGHT(-/-)) mice using gene targeting. Disruption of LIGHT significantly reduced CD8(+) T cell-cycle progression, leading to reduced proliferation to anti-CD3, anti-CD3/anti-CD28 or allogeneic stimulation, whereas proliferation of CD4(+) T cells remained unchanged. In contrast to the observed proliferative defects, isolated CD8(+) T cells from LIGHT(-/-) mice displayed normal cytotoxic effector function development when compared to wild-type CD8(+) T cells. Underlying a potential mechanism of reduced CD8(+) T cell proliferation, LIGHT(-/-) CD8(+) T cells displayed reduced surface levels of CD25 and a diminished ability to proliferate in response to exogenous IL-2. Furthermore, addition of IL-12 to LIGHT(-/-) CD8(+) T cell cultures could not ameliorate this proliferative defect. These results reveal a potential mechanism of action for LIGHT as a positive regulator of CD8(+) T cell expansion, but not lytic effector function development.


Subject(s)
CD8-Positive T-Lymphocytes/immunology , Lymphocyte Activation , Membrane Proteins/physiology , Tumor Necrosis Factor-alpha/physiology , Animals , Apoptosis , Genetic Vectors , Interleukin-12/pharmacology , Interleukin-2/pharmacology , Membrane Proteins/genetics , Mice , Mice, Inbred BALB C , Mice, Knockout , Receptors, Interleukin-2/metabolism , T-Lymphocyte Subsets/immunology , T-Lymphocytes, Cytotoxic/immunology , Tumor Necrosis Factor Ligand Superfamily Member 14 , Tumor Necrosis Factor-alpha/genetics , Tumor Necrosis Factor-alpha/immunology
6.
Mol Vis ; 8: 407-15, 2002 Oct 31.
Article in English | MEDLINE | ID: mdl-12432342

ABSTRACT

PURPOSE: To study the role of mimecan, a member of the small leucine-rich proteoglycans (SLRPs) gene family and one of the major components of the cornea and other connective tissues, mice that lack a functional mimecan gene were generated and characterized. METHODS: Mimecan-deficient mice were generated by gene-targeting using standard techniques. Mice were genotyped by Southern blot analysis. The absence of mimecan transcripts was confirmed by Northern blot analysis. Corneal clarity was examined by slit lamp biomicroscopy. The strength of the skin was evaluated using a biomechanical skin fragility test. Collagen morphology in cornea and skin preparations from mimecan-null and control wild-type mice was analyzed by transmission electron microscopy. The diameter of collagen fibrils in these tissues was determined by morphometric analysis. RESULTS: Mice lacking mimecan appear to develop normally, are viable and fertile. In a controlled laboratory environment they do not display an evident pathological phenotype compared to wild type mice. Examination of corneal clarity and measurements of corneal thickness show no significant changes in the cornea. However, a skin fragility test revealed a moderate reduction in the tensile strength of skin from mutant mice. Ultrastructural analyses show, on average, thicker collagen fibrils in both corneal and skin preparations from mimecan-null mice. Collagen fibrils from the cornea of mutant mice show an average diameter of 31.84+/-0.322 nm, versus 22.40+/-0.296 nm in their wild type litter-mates. The most pronounced increase in collagen fibril diameter was found in the skin of mimecan-null mice, who demonstrated an average diameter of 130.33+/-1.769 nm, versus 78.82+/-1.157 nm in the wild type mice. In addition, size variability and altered collagen morphology was detected in dorsal and tail skin preparations from the mutant mice. CONCLUSIONS: The results of the present study demonstrate that mimecan, similar to other members of the SLRP gene family, has a role in regulating collagen fibrillogenesis in vivo. Further studies, such as functional challenges, an evaluation of potential compensation by other proteins (including members of the SLRP family), and generation of double-knockouts will be necessary to fully uncover physiological functions of mimecan in mice.


Subject(s)
Cornea/metabolism , Fibrillar Collagens/metabolism , Glycoproteins/physiology , Skin/metabolism , Animals , Blotting, Northern , Blotting, Southern , Bone and Bones/diagnostic imaging , Cornea/pathology , Cornea/ultrastructure , Female , Fibrillar Collagens/ultrastructure , Growth Substances/physiology , Intercellular Signaling Peptides and Proteins , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Radiography , Skin/pathology , Skin/ultrastructure
7.
Comp Med ; 52(5): 456-60, 2002 Oct.
Article in English | MEDLINE | ID: mdl-12405640

ABSTRACT

Elimination of an enzootic infection of mouse hepatitis virus (MHV) from a large population of genetically engineered mice was accomplished by selecting seropositive, non-infective breeders for a newly restored MHV-free breeding colony. An ELISA was used to test for the presence of MHV-specific antibody, and TaqMan reverse transcriptase-polymerase chain reaction (RT-PCR) analysis was used to detect MHV in the feces. After 10 weeks of intentional exposure, approximately 30% of mice with MHV antibodies continued to shed MHV in the feces. A natural transmission study was conducted to validate that positive fecal RT-PCR results indicated presence of infective virus. Sentinel results from the re-instituted breeding colony indicated that MHV was successfully eliminated by use of RT-PCR analysis for selection of non-infective mice.


Subject(s)
Disease Outbreaks/veterinary , Hepatitis, Viral, Animal/diagnosis , Murine hepatitis virus/enzymology , Reverse Transcriptase Polymerase Chain Reaction/veterinary , Taq Polymerase/metabolism , Animals , Antibodies, Viral/immunology , Breeding , Disease Outbreaks/prevention & control , Enzyme-Linked Immunosorbent Assay/veterinary , Feces/virology , Female , Hepatitis, Viral, Animal/prevention & control , Hepatitis, Viral, Animal/transmission , Male , Mice , Murine hepatitis virus/genetics , Murine hepatitis virus/immunology , Murine hepatitis virus/isolation & purification , RNA, Messenger/analysis , RNA, Viral/analysis , Reverse Transcriptase Polymerase Chain Reaction/methods , Taq Polymerase/genetics
8.
Endocrinology ; 143(10): 3994-4006, 2002 Oct.
Article in English | MEDLINE | ID: mdl-12239111

ABSTRACT

Vasoactive intestinal polypeptide (VIP) and pituitary adenylate cyclase-activating peptide (PACAP) are two closely related peptides that bind two homologous G protein-coupled receptors, VIP/PACAP receptor 1 (VPAC1R) and VIP/PACAP receptor II (VPAC2R), with equally high affinity. Recent reports suggest that VPAC2R plays a role in circadian rhythm and T cell functions. To further elucidate the functional activities of VPAC2R, we generated VPAC2R-deficient mice by deleting exons VIII-X of the VPAC2R gene. The VPAC2R-deficient mice showed retarded growth and had reduced serum IGF-I levels compared with gender-matched, wild-type siblings. The mutant mice appeared healthy and fertile at a young adult age. However, older male mutant mice exhibited diffuse seminiferous tubular degeneration with hypospermia and reduced fertility rate. The mutant mice appeared to have an increase in insulin sensitivity. VPAC2R-deficient mice had increased lean mass and decreased fat mass with reduced serum leptin levels. Indirect calorimetry experiments showed that the respiratory quotient values immediately following the transition into the dark cycle were significantly higher in male knockout mice for about 4 h. Additionally, male and female VPAC2R-deficient mice presented an increased basal metabolic rate (23% and 10%, respectively) compared with their wild-type siblings. Our results suggest that VPAC2R plays an important role in growth, basal energy expenditure, and male reproductive functions.


Subject(s)
Basal Metabolism/physiology , Growth/physiology , Receptors, Vasoactive Intestinal Peptide/physiology , Amino Acid Sequence/genetics , Animals , Body Composition , Female , Growth Disorders/genetics , Infertility, Male/genetics , Insulin/physiology , Insulin-Like Growth Factor I/analysis , Leptin/blood , Male , Mice , Mice, Knockout/genetics , Molecular Sequence Data , Receptors, Vasoactive Intestinal Peptide/deficiency , Receptors, Vasoactive Intestinal Peptide/genetics , Receptors, Vasoactive Intestinal Peptide, Type II , Reference Values , Seminiferous Tubules/pathology , Sex Characteristics , Sperm Count
9.
Endocrinology ; 143(7): 2469-77, 2002 Jul.
Article in English | MEDLINE | ID: mdl-12072376

ABSTRACT

The hypothalamic neuropeptide melanin-concentrating hormone (MCH) has been implicated in a variety of physiological functions including the regulation of feeding and energy homeostasis. Two MCH receptors (MCHR1 and MCHR2) have been identified so far. To decipher the functional role of the MCH receptors, we have generated and phenotypically characterized mice rendered deficient in MCHR1 expression by homologous recombination. Inactivation of MCHR1 results in mice (MCHR1-/-) that are resistant to diet-induced obesity. With a high-fat diet, body fat mass is significantly lower in both male (4.7 +/- 0.6 g vs. 9.6 +/- 1.2 g) and female (3.9 +/- 0.2 vs. 5.8 +/- 0.5 g) MCHR1-/- mice than that of the wild-type control (P < 0.01), but the lean mass remains constant. When normalized to body weight, female mice are hyperphagic, and male mice are hyperphagic and hypermetabolic, compared with wild-type mice. Consistent with the lower fat mass, both leptin and insulin levels are significantly lower in male MCHR1-/- mice than in the wild-type controls. Our data firmly establish MCHR1 as a mediator of MCH effects on energy homeostasis and suggest that inactivation of MCHR1 alone is capable to counterbalance obesity induced by a high-fat diet.


Subject(s)
Diet , Hyperphagia/genetics , Hyperphagia/psychology , Hypothalamic Hormones/physiology , Melanins/physiology , Obesity/genetics , Pituitary Hormones/physiology , Receptors, Pituitary Hormone/genetics , Receptors, Pituitary Hormone/physiology , Adipose Tissue/physiology , Animals , Basal Metabolism/drug effects , Basal Metabolism/genetics , Blotting, Northern , Blotting, Southern , Body Weight/genetics , Body Weight/physiology , Calorimetry, Indirect , DNA, Complementary/genetics , Dietary Fats/pharmacology , Energy Metabolism/genetics , Energy Metabolism/physiology , Female , Genotype , Male , Mice , Mice, Inbred Strains , Mice, Knockout , Obesity/physiopathology , Plasmids/genetics , Reverse Transcriptase Polymerase Chain Reaction , Sex Characteristics
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