Biomedical subjects

David J Baylink

Publications and source records attributed to David J Baylink.

At least 55 records · Page 3Linked to original sources

PubMed2003

Expression of a structurally unique osteoclastic protein-tyrosine phosphatase is driven by an alternative intronic, cell type-specific promoter.

An osteoclastic protein-tyrosine phosphatase (PTP-oc), essential for osteoclast activity, shows sequence identity with the intracellular domain of GLEPP1, a renal receptor-like transmembrane PTP. PTP-oc has been assumed to be a truncated variant of GLEPP1, resulting from alternative splicing. However, the 5'-untranslated region sequence of PTP-oc mRNA contains 217 bp from an intron of GLEPP1. There are no splicing acceptor sites at the PTP-oc transcription site. The intronic sequence flanking the 5' end of the PTP-oc transcription start site contains potential promoter elements essential for transcriptional initiation. To test the hypothesis that the PTP-oc gene has an alternative, tissue-specific, intronic promoter, the promoter activity of a 1.3-kb PCR fragment covering the 5'-flanking region of the PTP-oc gene was measured. The putative PTP-oc promoter fragment showed strong promoter activity in U937 cells. Mutation of the putative TATA box within the PTP-oc promoter abolished 60-90% of its promoter activity. The PTP-oc promoter fragment showed strong promoter activity in cells that express PTP-oc (U937 cells and RAW264.7 cells) but not in cells that do not express the enzyme (skin fibroblasts, TE85 cells, and HEK293 cells). These findings strongly support the conclusion that the 1.3-kb intronic fragment contains the tissue-specific, PTP-oc proximal promoter. Deletion and functional analyses indicate that the proximal 5' sequence flanking the TATA box of the PTP-oc contains potential repressor elements. The removal of the putative repressor elements led to the apparent loss of tissue specificity. In summary, we conclude that an intronic promoter within the GLEPP1 gene drives the expression of the PTP-oc in a cell type-specific manner. This GLEPP1/PTP-oc gene system is one of the very few systems in which two important tissue-specific enzymes are derived from the same gene by the use of alternative intronic promoters.

Alternative Splicing↗

PubMed2003

Lack of insulin-like growth factor I exaggerates the effect of calcium deficiency on bone accretion in mice.

Recent studies provide evidence that the GH/IGF-I axis plays a critical role in the regulation of bone accretion that occurs during puberty and that the peak bone mineral density (BMD) is dependent on the amount of dietary calcium intake during the active growth phases. To evaluate whether IGF-I deficiency exaggerates the effect of calcium deficiency on bone accretion during active growth phases, IGF-I knockout (KO) and wild-type (WT) mice were fed with low calcium (0.01%) or normal calcium (0.6%) for 2 wk during the pubertal growth phase and were labeled with tetracycline. The low calcium diet caused significant decreases in endosteal bone formation parameters and a much greater increase in the resorbing surface of both the endosteum and periosteum of the tibia of IGF-I KO mice compared with WT mice. Accordingly, femur BMD measured by dual energy x-ray absorptiometry or peripheral quantitative computed tomography increased significantly in IGF-I WT mice fed the low calcium diet, but not in IGF-I KO mice. IGF-I-deficient mice fed the normal calcium diet showed elevated PTH levels, decreased serum 1,25-dihydroxyvitamin D and serum calcium levels at baseline. Serum calcium changes due to calcium deficiency were greater in IGF-I KO mice compared with WT mice. PTH levels were 7-fold higher in IGF-I KO mice fed normal calcium compared with WT mice, which was further elevated in mice fed the low calcium diet. Treatment of IGF-I-deficient lit/lit mice with GH decreased the serum PTH level by 70% (P < 0.01). Based on these and past findings, we conclude that: 1) IGF-I deficiency exaggerates the negative effects of calcium deficiency on bone accretion; and 2) IGF-I deficiency may lead to 1,25-dihydroxyvitamin D deficiency and elevated PTH levels even under normal calcium diet.

Absorptiometry, Photon↗

PubMed2003

Analytical and clinical evaluation of the Bio-Rad HPLC kit for measurement of type I collagen cross links.

The measurement of hydroxylysylpyridinoline (PYD) and lysylpyridinoline (DPD), the degradation products of type I collagen, by manual HPLC assay posed practical difficulties. The present study was undertaken to evaluate the first commercially available HPLC kit, which provides a convenient substitute for cumbersome classical HPLC methods. The HPLC procedure is based on an improved sample clean-up chromatography, convenient ready-to-use HPLC reagents, and quicker isocratic elution of PYR and DPD on reverse-phase analytical column. The analytical parameters assessed include sensitivity, within- and between-assay variation, method comparison, recoveries, and interference. Clinical evaluation included discriminatory power of PYD and DPD and response to treatment of osteoporosis patients with Alendronate. DPD and PYD concentrations showed linear ( r(2) > 0.99) response between 10-400 pmol/ml and 75-4000 pmol/ml, respectively. The average within-assay imprecision, over a range of clinically relevant cross-links concentrations, was CV < 7% ( n = 30). The total imprecision ( n = 35 days), by ANOVA, for PYD and DPD was CV < 7.5% and CV < 10%, respectively. Average spike recovery was 95.4% +/- 6.5%. Comparison with the historical HPLC method exhibited a close correlation ( r values between 0.87 and 0.91, P < 0.0001). Creatinine-corrected DPD in postmenopausal ( Z score = 2.4, P < 0.05, n = 17) and osteoporotic ( Z score = 3.0, P < 0.01, n = 29) women were 44% and 64% higher, respectively, compared to premenopausal samples ( n = 15). Similarly, PYD concentration was 26% and 54% higher in postmenopausal and postmenopausal osteoporotic women, respectively. There was a 47% ( P < 0.001) decrease in DPD concentration ( n = 16), and a 30% decrease in PYD concentration after 90 days of treatment of osteoporotic patients with Alendronate. DPD concentration correlated with N-telopeptide with an r value of 0.69 ( n = 67, P < 0.0001). The reported kit method is substantially simpler and precise than the manual method. DPD concentrations determined by the current method reaffirm the clinical value in identifying increased bone resorption in pathological conditions and monitoring response to antiresorptive therapy.

Adult↗

PubMed2003

Fluid flow shear stress stimulates human osteoblast proliferation and differentiation through multiple interacting and competing signal transduction pathways.

This study sought to assess the role of several signaling pathways in the fluid flow shear stress-induced proliferation and differentiation of normal human osteoblasts. We evaluated the effects of an effective dose of selective inhibitors of the extracellular signal-regulated kinases (ERK) pathway (PD98059 and U0126), the nitric oxide synthase pathway (N(omega)-nitro-L-arginine methyl ester), the cyclo-oxygenase pathway (indomethacin), or the Gi/o pathway (pertussis toxin [PTX]) on the flow-mediated effects. A 30-min steady flow shear stress at 20 dynes/cm(2) increased significantly [(3)H]thymidine incorporation (an indicator of proliferation), alkaline phosphatase activity (an index of osteoblast differentiation), phosphorylation of ERK, and expression of integrin beta1. PD98059, U0126, and N(omega)-nitro-L-arginine methyl ester completely blocked the shear stress-induced increases in ERK phosphorylation, [(3)H]thymidine incorporation, and alkaline phosphatase, but without an effect on integrin beta1 expression, indicating that the ERK and nitric oxide synthase pathways are essential for the shear stress-induced proliferation and differentiation of normal human osteoblasts and that each involves ERK activation but not integrin beta1 upregulation. Indomethacin blocked the shear stress-induced osteoblast proliferation and differentiation and integrin beta1 upregulation but not ERK activation, suggesting that the cyclo-oxygenase pathway (i.e., prostacyclin and/or prostaglandin E(2)) mediates the shear stress-induced osteoblast proliferation in an ERK-independent manner. In contrast, PTX completely blocked the flow-induced increase in integrin beta1 expression but had no effect on the increase in the ERK phosphorylation or [(3)H]thymidine incorporation. PTX not only did not inhibit but also significantly enhanced the stimulatory effect of shear stress on alkaline phosphatase activity, suggesting that a PTX-sensitive signaling pathway may have an inhibitory role in osteoblast differentiation. In summary, this study shows, for the first time, that the signal transduction mechanism of shear stress in osteoblasts is complex and involves multiple ERK-dependent and independent pathways, and provides circumstantial evidence that there may be a PTX-sensitive pathway that has completing effects with an unknown pathway on the differentiation of normal human osteoblasts.

Alkaline Phosphatase↗

PubMed2003

In vivo bone formation in fracture repair induced by direct retroviral-based gene therapy with bone morphogenetic protein-4.

This study sought to develop an in vivo gene therapy to accelerate the repair of bone fractures. In vivo administration of an engineered viral vector to promote fracture healing represents a potential high-efficacy, low-risk procedure. We selected a murine leukemia virus (MLV)-based retroviral vector, because this vector would be expected to target transgene expression to the proliferating periosteal cells arising shortly after bone fracture. This vector transduced a hybrid gene that consisted of a bone morphogenetic protein (BMP)-4 transgene with the BMP-2 secretory signal to enhance the secretion of mature BMP-4. The MLV vector expressing this BMP-2/4 hybrid gene or beta-galactosidase control gene was administered at the lateral side of the fracture periosteum at 1 day after fracture in the rat femoral fracture model. X-ray examination by radiograph and peripheral quantitative computed tomography at 7, 14, and 28 days after fracture revealed a highly significant enhancement of fracture tissue size in the MLV-BMP-2/4-treated fractures compared to the control fractures. The tissue was extensively ossified at 14 and 28 days, and the newly formed bone exhibited normal bone histology. This tissue also exhibited strong immunohistochemical staining of BMP-4. Additional control and MLV-BMP-2/4-treated animals each were monitored for 70 days to determine the fate of the markedly enhanced fracture callus. Radiographs showed that the hard callus had been remodeled and substantial healing at the fracture site had occurred, suggesting that the union of the bone at the fracture site was at least as high in the BMP-4-treated bone as in the control bone. There was no evidence of viral vector infection of extraskeletal tissues, suggesting that this in vivo gene therapy for fracture repair is safe. In summary, we have demonstrated for the first time that a MLV-based retroviral vector is a safe and effective means of introducing a transgene to a fracture site and that this procedure caused an enormous augmentation of fracture bone formation.

Animals↗

PubMed2003

Evidence that sensitivity to growth hormone (GH) is growth period and tissue type dependent: studies in GH-deficient lit/lit mice.

We previously found that the magnitude of skeletal deficits caused by GH deficiency varied during different growth periods. To test the hypothesis that the sensitivity to GH is growth period dependent, we treated GH-deficient lit/lit mice with GH (4 mg/kg body weight.d) or vehicle during the prepubertal and pubertal (d 7-34), pubertal (d 23-34), postpubertal (d 42-55), and adult (d 204-217) periods and evaluated GH effects on the musculoskeletal system by dual energy x-ray absorptiometry (DEXA) and peripheral quantitative computed tomography. GH treatment during different periods significantly increased total body bone mineral content, bone mineral density (BMD), bone area, and lean body mass and decreased percentage of fat compared with vehicle; however, the magnitude of change varied markedly depending on the treatment period. For example, the increase in total body BMD was significantly (P < 0.01) greater when GH was administered between d 42-55 (15%) compared with pubertal (8%) or adult (7.7%) periods, whereas the net loss in percentage of body fat was greatest (-56%) when GH was administered between d 204 and 216 and least (-27%) when GH was administered between d 7 and 35. To determine whether GH-induced anabolic effects on the musculoskeletal system are maintained after GH withdrawal, we performed DEXA measurements 3-7 wk after stopping GH treatment. The increases in total body bone mineral content, BMD, and lean body mass, but not the decrease in body fat, were sustained after GH withdrawal. Our findings demonstrate that the sensitivity to GH in target tissues is growth period and tissue type dependent and that continuous GH treatment is necessary to maintain body fat loss but not BMD gain during a 3-7 wk follow-up.

Absorptiometry, Photon↗

PubMed2003

Insulin-like growth factor regulates peak bone mineral density in mice by both growth hormone-dependent and -independent mechanisms.

To evaluate the relative contribution of the GH/IGF axis to the development of peak bone mineral density (BMD), we measured skeletal changes in IGF-I knockout (KO), IGF-II KO, and GH-deficient lit/lit mice and their corresponding control mice at d 23 (prepubertal), 31 (pubertal), and 56 (postpubertal) in the entire femur by dual energy x-ray absorptiometry and in the mid-diaphysis by peripheral quantitative computed tomography. Lack of growth factors resulted in different degrees of failure of skeletal growth depending on the growth period and the growth factor involved. At d 23, femoral length, size, and BMD were reduced by 25-40%, 15-17%, and 8-10%, respectively, in mice deficient in IGF-I, IGF-II, and GH compared with the control mice. During puberty, BMD increased by 40% in control mice and by 15% in IGF-II KO and GH-deficient mice, whereas it did not increase in the IGF-I KO mice. Disruption of IGF-I, but not IGF-II, completely prevented the periosteal expansion that occurs during puberty, whereas it was reduced by 50% in GH-deficient mice. At d 56, femoral length, size, and BMD were reduced by 40-55%, 11-18%, and 25-32%, respectively, in mice deficient in IGF-I, IGF-II, and GH compared with the control mice. Our data demonstrate that: 1) mice deficient in IGF-I exhibit a greater impairment in bone accretion than mice deficient in IGF-II or GH; 2) GH/IGF-I, but not IGF-II, is critical for puberty-induced bone growth; and 3) IGF-I effects on bone accretion during prepuberty are mediated predominantly via mechanisms independent of GH, whereas during puberty they are mediated via both GH-dependent and GH-independent mechanisms.

Absorptiometry, Photon↗

PubMed2003

Mapping quantitative trait loci that influence serum insulin-like growth factor binding protein-5 levels in F2 mice (MRL/MpJ X SJL/J).

Recent studies using twins and inbred strains of mice reveal evidence for genetic mechanisms contributing to variation in circulating levels of IGF-I, IGF-II, and IGF binding protein (IGFBP)-3. To examine the hypothesis that serum IGFBP-5 levels have a strong heritable component, we intercrossed two inbred strains of mice, MRL/MpJ and SJL, which exhibit 79% difference in serum IGFBP-5 levels (554 +/- 68 vs. 309 +/- 51 ng/ml respectively, P < 0.001). A genome-wide scan was carried out using 137 polymorphic markers in 633 F2 female mice. Serum IGFBP-5 levels in the F2 progeny showed a normal distribution with an estimated heritability of 74%. Whole genome-wide scans for cosegregation of genetic marker data with high or low serum IGFBP-5 levels revealed six different quantitative trait loci (QTL) in chromosomes 1, 9 (two), 10, and 11 (two), which together explained 24% of F2 variance. Chromosome 11 QTL exhibited the highest LOD score (7.5). Based on the past findings that IGFBP-5 is an important bone formation stimulator, we predicted IGFBP-5 to contribute to bone mineral density variation in F2 mice. Accordingly, we found two of the six IGFBP-5 QTLs (Chrs 1 and 11) identified for serum IGFBP-5 phenotype also showed significant association with total body bone mineral density phenotype (measured by dual energy x-ray absorptiometry) in the F2 mice.

Absorptiometry, Photon↗

PubMed2003

Patients with Van Buchem disease, an osteosclerotic genetic disease, have elevated bone formation markers, higher bone density, and greater derived polar moment of inertia than normal.

Van Buchem disease is an autosomal recessive disease characterized by overgrowth of the skeleton. In a group of Dutch patients the disease is thought to be due to a 52-kb deletion that results in decreased expression of the SOST gene. To further characterize the disease, the morphology of the metacarpals of six adult subjects and two juveniles with Van Buchem disease were measured on hand x-rays along with nine normal adults and nine adult carriers of the disease. Serum bone formation markers, alkaline phosphatase, type I procollagen peptide, and osteocalcin, and the urinary bone resorption marker, cross-linked N-telopeptide, were determined. Van Buchem patients had increased metacarpal outer diameter, inner diameter, cortical thickness, and bone mineral density. Calculated bone volume and derived polar moment of inertia were markedly elevated (elevations of 158 +/- 33% and 497 +/- 95%, respectively) consistent with increased bone strength. Serum procollagen peptide and osteocalcin were significantly higher in Van Buchem patients. Urinary cross-linked N-telopeptide was significantly elevated in Van Buchem patients. None of these changes was found in Van Buchem carriers. These observations indicate that decreased expression of the SOST gene can lead to increased bone formation and to stronger bones.

Adolescent↗

PubMed2003

Osteoblastic tartrate-resistant acid phosphatase: its potential role in the molecular mechanism of osteogenic action of fluoride.

Although type 5 TRACP is recognized as a histochemical and biochemical marker of osteoclasts, there is evidence that bone forming cells, osteoblasts, and osteocytes also express a type 5 TRACP. Accordingly, an osteoblastic type 5 TRACP has been purified from human osteoblasts and from bovine cortical bone matrices. Comparison of biochemical properties of osteoblastic type 5 TRACP with those of osteoclastic type 5 TRACP suggests that osteoblastic type 5 TRACP is a different isoenzyme from osteoclastic type 5 TRACP. Two properties of osteoblastic type 5 TRACP may be relevant to its physiological functions: (1) it acts as a protein-tyrosine phosphatase (protein tyrosine phosphorylation) under physiologically relevant conditions, and (2) it is sensitive to inhibition by clinically relevant concentrations of fluoride. Because fluoride is a stimulator of osteoblastic proliferation and differentiation and a potent osteogenic agent and because protein tyrosine phosphorylation plays an important regulatory role in cell proliferation and differentiation, these unique properties and other evidence summarized in this review led to the proposal that the osteogenic action of fluoride is mediated, at least in part, by the fluoride-mediated inhibition of osteoblastic type 5 TRACP/protein tyrosine phosphorylation, which leads to a stimulation of osteoblast proliferation and differentiation, and subsequently, an increase in bone formation.

Acid Phosphatase↗

PubMed2003

Congenic strains of mice for verification and genetic decomposition of quantitative trait loci for femoral bone mineral density.

Peak femoral volumetric bone mineral density (femoral bone mineral density) in C57BL/6J (B6) 4-month-old female mice is 50% lower than in C3H/HeJ (C3H) and 34% lower than in CAST/EiJ (CAST) females. Genome-wide analyses of (B6 x C3H)F2 and (B6 x CAST)F2 4-month-old female progeny demonstrated that peak femoral bone mineral density is a complex quantitative trait associated with genetic loci (QTL) on numerous chromosomes (Chrs) and with trait heritabilities of 83% (C3H) and 57% (CAST). To test the effect of each QTL on femoral bone mineral density, two sets of loci (six each from C3H and CAST) were selected to make congenic strains by repeated backcrossing of donor mice carrying a given QTL-containing chromosomal region to recipient mice of the B6 progenitor strain. At the N6F1 generation, each B6.C3H and B6.CAST congenic strain (statistically 98% B6-like in genomic composition) was intercrossed to obtain N6F2 progeny for testing the effect of each QTL on femoral bone mineral density. In addition, the femoral bone mineral density QTL region on Chr 1 of C3H was selected for congenic subline development to facilitate fine mapping of this strong femoral bone mineral density locus. In 11 of 12 congenic strains, 6 B6.C3H and 5 B6.CAST, femoral bone mineral density in mice carrying c3h or cast alleles in the QTL regions was significantly different from that of littermates carrying b6 alleles. Differences also were observed in body weight, femoral length, and mid-diaphyseal periosteal circumference among these 11 congenic strains when compared with control littermates; however, these latter three phenotypes were not consistently correlated with femoral bone mineral density. Analyses of eight sublines derived from the B6.C3H-1T congenic region revealed two QTLs: one located between 36.9 and 49.7 centiMorgans (cM) and the other located between 73.2 and 100.0 cM distal to the centromere. In conclusion, these congenic strains provide proof of principle that many QTLs identified in the F2 analyses for femoral bone mineral density exert independent effects when transferred and expressed in a common genetic background. Furthermore, significant differences in femoral bone mineral density among the congenic strains were not consistently accompanied by changes in body weight, femur length, or periosteal circumference. Finally, decomposition of QTL regions by congenic sublines can reveal additional loci for phenotypes assigned to a QTL region and can markedly refine genomic locations of quantitative trait loci, providing the opportunity for candidate gene testing.

Alleles↗

PubMed2003

A missense mutation in the mouse Col2a1 gene causes spondyloepiphyseal dysplasia congenita, hearing loss, and retinoschisis.

UNLABELLED: A missense mutation in the mouse Col2a1 gene has been discovered, resulting in a mouse phenotype with similarities to human spondyloepiphyseal dysplasia (SED) congenita. In addition, SED patients have been identified with a similar molecular mutation in human COL2A1. This mouse model offers a useful tool for molecular and biological studies of bone development and pathology. INTRODUCTION: A new mouse autosomal recessive mutation has been discovered and named spondyloepiphyseal dysplasia congenita (gene symbol sedc). MATERIALS AND METHODS: Homozygous sedc mice can be identified at birth by their small size and shortened trunk. Adults have shortened noses, dysplastic vertebrae, femora, and tibias, plus retinoschisis and hearing loss. The mutation was mapped to Chr15, and Col2a1 was identified as a candidate gene. RESULTS: Sequence analyses revealed that the affected gene is Col2a1, which has a missense mutation at exon 48 causing an amino acid change of arginine to cysteine at position 1417. Two human patients with spondyloepiphyseal dysplasia (SED) congenita have been reported with the same amino acid substitution at position 789 in the human COL2A1 gene. CONCLUSIONS: Thus, sedc/sedc mice provide a valuable model of human SED congenita with molecular and phenotypic homology. Further biochemical analyses, molecular modeling, and cell culture studies using sedc/sedc mice could provide insight into mechanisms of skeletal development dependent on Col2a1 and its role in fibril formation and cartilage template organization.

Amino Acid Sequence↗

PubMed2002

ADAM-9 is an insulin-like growth factor binding protein-5 protease produced and secreted by human osteoblasts.

IGF binding protein-5 (BP-5) is an important bone formation regulator. Therefore, elucidation of the identity of IGF binding protein-5 (BP-5) protease produced by osteoblasts is important for our understanding of the molecular pathways that control the action of BP-5. In this regard, BP-5 protease purified by various chromatographic steps from a conditioned medium of U2 human osteosarcoma cells migrated as a single major band, which comigrated with the protease activity in native PAGE and yielded multiple bands in SDS-PAGE under reducing conditions. N-Terminal sequencing of these bands revealed that three of the bands yielded amino acid sequences that were identical to that of alpha2 macroglobulin (alpha2M). Although alpha2M was produced by human osteoblasts (OBs), it was not found to be a BP-5 protease. Because alpha2M had been shown to complex with ADAM proteases and because ADAM-12 was found to cleave BP-3 and BP-5, we evaluated if one of the members of ADAM family was the BP-5 protease. On the basis of the findings that (1) purified preparations of BP-5 protease from U2 cell CM contained ADAM-9, (2) ADAM-9 is produced and secreted in high abundance by various human OB cell types, (3) purified ADAM-9 cleaved BP-5 effectively while it did not cleave other IGFBPs or did so with less potency, and (4) purified ADAM-9 bound to alpha2M, we conclude that ADAM-9 is a BP-5 protease produced by human OBs.

ADAM Proteins↗

PubMed2002

Construction of a Tc1-like transposon Sleeping Beauty-based gene transfer plasmid vector for generation of stable transgenic mammalian cell clones.

We have constructed a single plasmid-, Tc1-like transposon-based gene transfer vector, termed the Prince Charming vector (pPC). The pPC vector was constructed by ligating the CMV-driven "Sleeping Beauty" transposase gene downstream to the Tc1-like transposon inverted repeat (IR) elements and by inserting the RSV promoter (to drive expression of the gene-of-interest) along with a multiple cloning site (MCS), a polyadenylation signal, and the SV40 promoter-driven neomycin gene, at a site flanked by the transposon IR elements. To assess the utility of the pPC vector, we cloned a red fluorescent protein (RFP) gene into the pPC vector at the MCS and transfected human TE85 osteosarcoma cells with the pPC-RFP expression vector using Effectene. Stable transgenic cell clones expressing RFP were selected with G418 sulfate and individual clones were isolated. After 4 weeks of clonal isolation and expansion, 99% of cells in each randomly selected clone expressed RFP strongly. Aliquots of each clone were then maintained in either the presence or the absence of G418 sulfate and were passaged weekly. Even after 6 months in culture in the absence of G418 sulfate, approximately 90% of the cells in each clone still maintained a strong expression level of RFP, indicating that these transgenic cell clones were stable and that the clonal stability of these clones did not require a constant selection pressure. In conclusion, we have developed a single plasmid-, Tc1-like transposon-based gene transfer vector that can be used to generate stable transgenic mammalian cell clones.

Chromosome Inversion↗

PubMed2002

Quantitative trait loci that harbor genes regulating muscle size in (MRL/MPJ x SJL/J) F(2) mice.

The genetic mechanisms that determine muscle size have not been elucidated, even though it is a key musculoskeletal parameter that reflects muscle strength. In this study, we performed a high-density genome-wide scan using 633 (MRL/MPJ x SJL/J) F(2) intercross 7-week-old mice to identify quantitative trait loci (QTL) involved in the determination of muscle size. Significant QTL were identified for muscle size and body length. Muscle size (adjusted by body length) QTL were identified on chromosomes 7, 9, 11, 14 (two QTL) and 17, which together explained 19.2% of phenotypic variance in F(2)mice, while body length QTL were located on chromosome 2 (two QTL), 9, 11 and 17 which accounted for 28.3% of phenotypic variance in F(2) mice. Three significant epistatic interactions between different QTL positions from muscle size and body length were identified ( P <0.01) on chromosomes 2, 9, 14 and 17, which explained 16.1% of the variance in F(2) mice.

Alleles↗

PubMed2002

Quantitative trait loci (QTL) for lean body mass and body length in MRL/MPJ and SJL/J F(2) mice.

Studies on the genetic mechanisms involved in the regulation of lean body mass (LBM) in mammals are minimal, although LBM is associated with a competent immune system and an overall good (healthy) body functional status. In this study, we performed a high-density genome-wide scan using 633 (MRL/MPJ x SJL/J) F(2) intercross to identify the quantitative trait loci (QTL) involved in the regulation of LBM. We hypothesized that additional QTL can be identified using a different mouse cross (MRL/SJL cross). Ten QTL were identified for LBM on chromosomes (chrs) 2, 6, 7, 9,13 and 14. Of those ten, QTL on chrs 6, 7 and 14 were exclusive to LBM, while QTL on chrs 4 and 11 were exclusively body length. LBM QTL on chrs 2 and 9 overlap with those of size. Altogether, the ten LBM QTL explained 41.2% of phenotypic variance in F(2) mice. Five significantly interacting loci that may be involved in the regulation of LBM were identified and accounted for 24.4% of phenotypic variance explained by the QTL. Five epistatic interactions, contributing 22.9% of phenotypic variance, were identified for body length. Interacting loci on chr 2 may influence LBM by regulating body length. Therefore, epistatic interactions as well as single QTL effects play an important role in the regulation of LBM.

Alleles↗

PubMed2002

Chromosomal regions harboring genes for the work to femur failure in mice.

The work to failure is defined as the maximum energy bone can absorb before breaking, and therefore is a direct test of the risk of fracture. To determine the genetic loci influencing work to failure, we have performed a high density genome-wide scan in 633 (MRL x SJL) F(2) female mice. Five loci ( P<0.005) with significant effects on work to failure were found on chromosomes 2, 7, 8, 9, and X, which collectively explained around 20% variance of work to femur failure in F(2) mice. Of those, only the QTL on chromosome 9 was concordant with bone mineral density (BMD) QTLs. Eight significant interactions ( P<0.01) between marker loci were identified, which accounted for an equivalent amount of F(2) variance (23%) to combined single QTL effects. Our results demonstrate that most of the genetic loci regulating work to failure are different from those for BMD in the 7-week-old female mice. If this is also true in humans, this finding will challenge the predictive value of BMD for the risk of fracture.

Animals↗

PubMed2002

Gene expression between a congenic strain that contains a quantitative trait locus of high bone density from CAST/EiJ and its wild-type strain C57BL/6J.

Peak bone density is an important determining factor of future osteoporosis risk. We previously identified a quantitative trait locus (QTL) that contributes significantly to high bone density on mouse chromosome 1 from a cross between C57BL/6J (B6) and CAST/EiJ (CAST) mouse strains. We then generated a congenic strain, B6.CAST-1T, in which the chromosomal fragment containing this QTL had been transferred from CAST to the B6 background. The congenic mice have a significantly higher bone density than the B6 mice. In this study we performed cDNA microarray analysis to evaluate the gene expression profile that might yield insights into the mechanisms controlling the high bone density by this QTL. This study led to several interesting observations. First, approximately 60% of 8,734 gene accessions on GEM I chips were expressed in the femur of B6 mice. The expression and function of two-thirds of these expressed genes and ESTs have not been documented previously. Second, expression levels of genes related to bone formation were lower in congenic than in B6 mice. These data are consistent with a low bone formation in the congenic mice, a possibility that is confirmed by reduced skeletal alkaline phosphatase activity in serum compared with B6 mice. Third, expression levels of genes that might have negative regulatory action on bone resorption were higher in congenic than in B6 mice. Together these findings suggest that the congenic mice might have a lower bone turnover rate than B6 mice and raise the possibility that the high bone density in the congenic mice could be due to reduced bone resorption rather than increased bone formation.

Animals↗