| 1. |
- Halling Linder, Cecilia, et al.
(författare)
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Glycosylation differences contribute to distinct catalytic properties among bone alkaline phosphatase isoforms.
- 2009
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Ingår i: Bone. - : Elsevier BV. - 1873-2763 .- 8756-3282. ; 45:5, s. 987-993
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Tidskriftsartikel (refereegranskat)abstract
- Three circulating human bone alkaline phosphatase (BALP) isoforms (B1, B2, and B/I) can be distinguished in healthy individuals and a fourth isoform (B1x) has been discovered in patients with chronic kidney disease and in bone tissue. The present study was designed to correlate differing glycosylation patterns of each BALP isoform with their catalytic activity towards presumptive physiological substrates and to compare those properties with two recombinant isoforms of the tissue-nonspecific ALP (TNALP) isozyme, i.e., TNALP-flag, used extensively for mutation analysis of hypophosphatasia mutations and sALP-FcD(10), a chimeric enzyme recently used as therapeutic drug in a mouse model of infantile hypophosphatasia. The BALP isoforms were prepared from human osteosarcoma (SaOS-2) cells and the kinetic properties were evaluated using the synthetic substrate p-nitrophenylphosphate (pNPP) at pH 7.4 and 9.8, and the three suggested endogenous physiological substrates, i.e., inorganic pyrophosphate (PP(i)), pyridoxal 5'-phosphate (PLP), and phosphoethanolamine (PEA) at pH 7.4. Qualitative glycosylation differences were also assessed by lectin binding and precipitation. The k(cat)/K(M) was higher for B2 for all the investigated substrates. The catalytic activity towards PEA was essentially undetectable. The kinetic activity for TNALP-flag and sALP-FcD(10) was similar to the activity of the human BALP isoforms. The BALP isoforms differed in their lectin binding properties and dose-dependent lectin precipitation, which also demonstrated differences between native and denatured BALP isoforms. The observed differences in lectin specificity were attributed to N-linked carbohydrates. In conclusion, we demonstrate significantly different catalytic properties among the BALP isoforms due to structural differences in posttranslational glycosylation. Our data also suggests that PEA is not an endogenous substrate for the BALP isoforms or for the recombinant TNALP isoforms. The TNALP-flag and the sALP-FcD(10) isoforms faithfully mimic the biological properties of the human BALP isoforms in vivo validating the use of these recombinant enzymes in studies aimed at dissecting the pathophysiology and treating hypophosphatasia.
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| 2. |
- Linder, Cecilia, et al.
(författare)
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Isozyme profile and tissue-origin of alkaline phosphatases in mouse serum
- 2013
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Ingår i: Bone. - : Elsevier. - 8756-3282 .- 1873-2763. ; 53:2, s. 399-408
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Tidskriftsartikel (refereegranskat)abstract
- Mouse serum alkaline phosphatase (ALP) is frequently measured and interpreted in mammalian bone research. However, little is known about the circulating ALPs in mice and their relation to human ALP isozymes and isoforms. Mouse ALP was extracted from liver, kidney, intestine, and bone from vertebra, femur and calvaria tissues. Serum from mixed strains of wild-type (WT) mice and from individual ALP knockout strains were investigated, i.e., Alpl(-/-) (a.k.a. Akp2 encoding tissue-nonspecific ALP or TNALP), Akp3(-/-) (encoding duodenum-specific intestinal ALP or dIALP), and Alpi(-/-) (a.k.a. Akp6 encoding global intestinal ALP or gIALP). The ALP isozymes and isoforms were identified by various techniques and quantified by high-performance liquid chromatography. Results from the WT and knockout mouse models revealed identical bone-specific ALP isoforms (B/I. B1, and B2) as found in human serum, but in addition mouse serum contains the B1x isoform only detected earlier in patients with chronic kidney disease and in human bone tissue. The two murine intestinal isozymes, dIALP and gIALP, were also identified in mouse serum. All four bone-specific ALP isoforms (B/I, B1x, B1, and B2) were identified in mouse bones, in good correspondence with those found in human bones. All mouse tissues, except liver and colon, contained significant ALP activities. This is a notable difference as human liver contains vast amounts of ALP. Histochemical staining, Northern and Western blot analyses confirmed undetectable ALP expression in liver tissue. ALP activity staining showed some positive staining in the bile canaliculi for BALB/c and FVB/N WT mice, but not in C57BI/6 and ICR mice. Taken together, while the main source of ALP in human serum originates from bone and liver, and a small fraction from intestine (andlt;5%), mouse serum consists mostly of bone ALP, including all four isoforms, B/I, B1x, B1, and B2, and two intestinal ALP isozymes dIALP and gIALR We suggest that the genetic nomenclature for the Alpl gene in mice (i.e., ALP liver) should be reconsidered since murine liver has undetectable amounts of ALP activity. These findings should pave the way for the development of user-friendly assays measuring circulating bone-specific ALP in mouse models used in bone and mineral research.
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| 3. |
- Sharp, Christopher, et al.
(författare)
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Increased bone alkaline phosphatase levels do not necessarily cause hypermineralization per se
- 2016
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Ingår i: Bone. - : Elsevier. - 8756-3282 .- 1873-2763. ; 89, s. 83-84
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- Diminished bone health and increased risk of fracture is increasingly recognised as a complication of diabetes mellitus and even as a consequence of its management. We read with interest the recently published review by Starup-Linde and Vestergaard [1] about bone turnover markers (BTMs) in patients with types 1 and 2 diabetes mellitus and their value in assessing bone health in diabetic patients. In brief, 611 eligible studies were identified of which 47 were included: 1 meta-analysis, 29 cross-sectional, 13 randomized controlled trials and 4 longitudinal studies.
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| 4. |
- Swolin-Eide, Diana, 1968, et al.
(författare)
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Variation of bone acquisition during growth hormone treatment in children can be explained by proteomic biomarkers, bone formation markers, body composition and nutritional factors
- 2018
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Ingår i: Bone. - : Elsevier BV. - 1873-2763 .- 8756-3282. ; 116, s. 144-153
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Tidskriftsartikel (refereegranskat)abstract
- OBJECTIVE: Growth hormone (GH) regulates both longitudinal growth and bone acquisition in children, and has profound metabolic effects. The aim was to investigate the association between proteomic biomarkers, body fat, nutrition and bone formation markers, and longitudinal growth in response to GH during the first year of treatment. The degree to which changes in these factors could explain variations in GH-dependent longitudinal growth and bone mineralization was also assessed. METHODS: The individualized GH dose trial included 128 short prepubertal children with either normal (non-GH-deficient) or reduced levels of GH secretion (GH-deficient) (mean age+/-SD, 8.6+/-2.6years; 90 boys), i.e., with a broad range of GH-secretion and GH-responsiveness, receiving GH treatment (mean 43mug/kg/day). Blood samples were taken and dual-energy X-ray absorptiometry (DXA) measured at baseline and 1year of treatment. Step-wise multiple regression models were constructed including three steps with different independent variables added at each step to explain the variance in outcome variables (heightSDS, bone mineral content (BMC) and bone mineral density (BMD). Independent variables included in Step I were previously identified proteomic markers related to GH treatment response, bone formation markers (intact PINP, bone-specific alkaline phosphatase and osteocalcin), variables at treatment start (GH dose mU/kg/day, GH maximum secretion, and difference between child's current and mid-parental heightSDS). Step II explored the added influence of body composition data (body mass index or DXA). Step III explored the added influence of serum nutritional markers and hormones. RESULTS: Step I variables explained 71% of the variation in first year heightSDS gain, median (minimum-maximum) 0.8 (0.24-1.67); and the proportion explained rose to 73% following inclusion of step II variables and 75% following step III. Corresponding values for total body BMC were 58%, 78%, and 80%, respectively. Proportions fell by approximately 20% when BMC was adjusted for height; 33%, 57%, and 57% for steps I, II, and III, respectively. Corresponding values for total body BMD were 29%, 39%, and 45%, respectively. CONCLUSION: For total BMC, as much as 80% of the variation during the first year of GH treatment could be explained by proteomic biomarkers, body fat, nutrition and bone formation markers, whereas for height-adjusted BMC 57% could be explained. The inclusion of information about either body composition (fat/lean mass) or nutritional markers contributed with approximately 20%. The variation in heightSDS gain could be explained to 75%. Hence, information of fat or nutrition markers was needed for explaining the variation in bone acquisition to the same magnitude as explaining the variation in height response.
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| 5. |
- Tubić, Bojan, 1984, et al.
(författare)
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Different osteocalcin forms, markers of metabolic syndrome and anthropometric measures in children within the IDEFICS cohort
- 2016
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Ingår i: Bone. - : ELSEVIER SCIENCE INC. - 8756-3282 .- 1873-2763. ; 84, s. 230-236
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Tidskriftsartikel (refereegranskat)abstract
- Objective: Osteocalcin (OC), an aboundant non-collagenous bone protein, is inversely associated with parameters of glucose metabolism. Interactions between bone tissue and energy metabolism have not been thoroughly investigated during childhood. This study investigated OC, metabolic parameters and anthropometric characteristics in normal weight and overweight/obese children. Methods: This study comprised 108 (46 normal weight/62 overweight/obese) Swedish 2-9 year old children. Anthropometric data, insulin, glucose, glycosylated haemoglobin (HbA1c), HOMA index, vitamin D, adiponectin, total OC, carboxylated OC (cOC) and undercarboxylated OC (ucOC) were analysed. Results: No difference was found for total OC between the normal and overweight/obese groups, with a mean (+/- SD) value of 82.6 ( +/- 2.8) ng/mL and 77.0 ( +/- 2.4) ng/mL, (P = 0.11), respectively. Overweight children had lower cOC levels, mean 69.1 ( +/- 2.2) ng/mL, vs. normal weight children, mean 75.6 ( +/- 2.5) ng/mL (P = 0.03). The mean ucOC levels of 7.9 ( +/- 0.4) ng/mL in overweight children did not differ vs. normal weight children, mean level 7.0 (+/- 0.4) ng/mL, (P = 0.067). None of the three OC forms correlated with any of the measured parameters. Conclusions: The cOC levels were lower in overweight children. There was no correlation between the three OC forms and any of the measured anthropometric or metabolic parameters. OC has been suggested to have a possible metabolic role, but in general the current study in prepubertal children does not support the hypothesis of an association between OC and a positive metabolic profile. (C) 2016 Elsevier Inc. All rights reserved.
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| 6. |
- Whyte, Michael P., et al.
(författare)
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Persistent idiopathic hyperphosphatasemia from bone alkaline phosphatase in a healthy boy
- 2020
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Ingår i: Bone. - : ELSEVIER SCIENCE INC. - 8756-3282 .- 1873-2763. ; 138
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Tidskriftsartikel (refereegranskat)abstract
- Alkaline phosphatase (ALP) in humans comprises a family of four cell-surface phosphomonoester phosphohydrolase isozymes. Three genes separately encode the "tissue-specific" ALPs whereas the fourth gene encodes ubiquitous homodimeric "tissue-nonspecific" ALP (TNSALP) richly expressed in bone, liver, kidney, and developing teeth. TNSALP monomers have five putative N-linked glycosylation sites where different post-translational modifications account for this isozymes distinctive physicochemical properties in different organs. Three bone-derived TNSALP (BALP) isoforms (B/I, B1, and B2) are present in healthy serum, whereas a fourth BALP isoform (Blx) can circulate in chronic kidney disease. Herein, we report a healthy boy with persistent hyperphosphatasemia due to BALP levels two- to threefold higher than age-appropriate reference values. Highperformance liquid chromatography, electrophoresis, heat inactivation, catalysis inhibition, and polyethylene glycol precipitation revealed increased serum B/I, B1, and B2 differing from patterns found in skeletal diseases. BA was similar to 23-fold elevated. Absence of mental retardation and physical stigmata excluded Mabry syndrome, the ALP-anchoring disorder causing hyperphosphatasemia. Routine biochemical studies indicated intact mineral homeostasis. Serum N-terminal propeptide of type I procollagen (P1NP) level was normal, but C-terminal crosslinking telopeptide of type I collagen (CTX) level was elevated. However, radiological studies showed no evidence for a generalized skeletal disturbance. Circulating pyridoxal 5-phosphate, a TNSALP natural substrate, was not low despite the laboratory hyperphosphatasemia, thereby suggesting BALP phosphohydrolase activity was not elevated endogenously. Mutation analysis of the ALPL gene encoding TNSALP revealed no defect. His non-consanguineous healthy parents had serum total ALP activity and BALP protein levels that were normal. Our patients sporadic idiopathic hyperphosphatasemia could reflect altered post-translational modification together with increased expression and/or impaired degradation of BALP.
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| 7. |
- Aspenberg, Per, et al.
(författare)
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Targeting RANKL for reduction of bone loss around unstable implants: OPG-Fc compared to alendronate in a model for mechanically induced loosening
- 2011
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Ingår i: BONE. - : Elsevier Science B.V., Amsterdam.. - 8756-3282. ; 48:2, s. 225-230
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Tidskriftsartikel (refereegranskat)abstract
- Orthopedic joint prostheses may loosen because of localized bone resorption. Despite initial optimism, there are no reports showing that bisphosphonates can stop the progression of prosthetic loosening once it has begun. This might be due to the strong resorptive stimulus, which continuously recruits new osteoclasts. Therefore, we hypothesized that a treatment targeting osteoclast recruitment would be more efficacious than a treatment reducing osteoclast activity. We used a previously described rat model for instability-induced bone resorption, and compared OPG-Fc with alendronate at a clinically relevant or an extreme dose. A titanium plate was osseointegrated at the rat tibial surface. Instability was simulated by a piston, moving perpendicularly to the bone surface. Piston movement induced bone loss via hydrostatic pressure or fluid flow. Rats were randomized to 5 groups (total n = 56), of which 4 were subjected to instability and one was stable. The unstable groups were injected with either high-dose OPG-Fc (10 mg/kg, twice weekly), a high dose of alendronate (20 mu g /kg/day), an extreme dose of alendronate (200 mu g/kg/day) or saline. Significant protection against resorption could only be shown for OPG-Fc and the extreme alendronate dose. Both alendronate doses reduced serum levels of tartrate-resistant acid phosphatase isoform 5b to a similar extent, demonstrating that the lower dose was able to reduce resorption in the normally remodeling skeleton, although not in the osteolytic lesions caused by instability. Osteoclast numbers in the lesion were increased by the lower bisphosphonate dose and reduced by OPG-Fc. The results suggest the possibility of targeting osteoclast recruitment via the RANKL system in patients with impending prosthetic loosening.
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| 8. |
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