Kits for classifying a subject having or at risk for developing idiopathic scoliosis
A method of classifying a subject having idiopathic scoliosis (IS) or at risk of developing IS comprising: determining the cellular response to Gi stimulation in a cell sample from the subject in the presence of OPN; determining the cellular response to Gi stimulation in a cell sample from the subject in the absence of OPN; and comparing the cellular response obtained in the presence of OPN with the cellular response obtained in the absence of OPN, whereby the comparing step enables the classification of the subject into one IS functional group. Also provided is the use of the foregoing method to classify borderline subjects and kits for applying the methods.
1. A kit for classifying a subject having idiopathic scoliosis (IS) or at risk of developing IS comprising:
(a) a purified recombinant osteopontin (rOPN) preparation; and
(b) a ligand for Gi stimulation;
wherein said kit optionally further comprises in addition to (a) and (b),
(c) a ligand for Gs stimulation; and/or
(d) one or more antibodies for detecting Giα phosphorylation.
2. The kit of claim 1 , wherein said ligand for Gi stimulation is selected from the group consisting of: (a) melatonin for the MT2 receptor; (b) 1-[3-(3,4-Methylenedioxyphenoxy)propyl]-4-phenyl-piperazine (BP554) maleate for the 5-HT1A receptor; (c) 5-bromo-N-(4,5-dihydro-IH-imidazol-2-yl)-6-quinoxalinamine (UK14304 ) for the a2-AD receptor; (d) 1-Deoxy-1-[6-[[3-iodophenyl)methyl]amino]-9H-purin-9-yl]-N-methyl-β-D-ribofuranuronamide (IB-MECA) for the A3 receptor; (e) Lysophosphatidic acid (LPA) for the LPA receptor; (f) (2S)-2-[[2-[[(2R)-2-[[(2S)-2-amino-3-(4-hydroxyphenyl)propanoyl]amino]propanoyl]amino]acetyl]-methylamino]-N-(2-hydroxyethyl)-3-phenylpropanamide (DMAGO) for the mu-opioid receptor; (g) 1-(6-Amino-9H-purin-9-yl)-1-deoxy-N-ethyl-β-D-ribofuranuronamide (NECA) for the adenosine receptors; (h) somatostatin for the SSTR receptor; (i) peptide MMK-1 for the FPR2 receptor; (j) Apelin-17 for the APJR receptor; (k) quinpirole for the D2 and D3 receptors; and (l) N-Cyclohexyl-7-chloro-1-[2-(4-morpholinyl)ethyl]quinolin-4(1H)-one-3-carboxamide (CB65) for the CB2 receptor.
3. The kit of claim 2 , comprising a ligand for Gs stimulation, wherein said ligand for Gs stimulation is a ligand for a GsPCR selected from the group consisting of: 5-HT4, 5-HT7, ACTH receptor, Adenosine receptor types A2a and A2b, Arginine vasopressin receptor 2, Calcitonin receptor, Calcitonin gene-related peptide receptor, corticotropin-releasing hormone receptor, Dopamine receptors D1-like family (D1 and D5), FSH-receptor, Gastric inhibitory polypeptide receptor, Glucagon receptor, Histamine H2 receptor, Luteinizing hormone/choriogonadotropin receptor, melanocortin receptor, MC1R, MC3R, MC4R, MC5R, Parathyroid hormone receptor 1, Prostaglandin receptor types D2 and I2, secretin receptor and thyrotropin receptor.
4. The kit of claim 3 , further comprising a reagent for measuring cellular cAMP concentration.
5. The kit of claim 2 , comprising a ligand for Gs stimulation, wherein said ligand for Gs stimulation is a ligand for a GsPCR selected from the group consisting of: β-adrenergic receptors types β1, β2 and β3.
6. The kit of claim 1 , wherein the ligand is a synthetic ligand.
7. The kit of claim 1 , comprising melatonin, iodomelatonin or phenylmelatonin as a ligand for Gi stimulation.
8. A composition for classifying a subject having idiopathic scoliosis (IS) comprising:
(a) a peripheral blood mononuclear cell (PBMC) sample from the subject;
(b) osteopontin (OPN); and
(c) a ligand for Gi stimulation;
wherein said composition optionally further comprises in addition to (a), (b) and (c),
(d) a ligand for Gs stimulation.
9. The composition of claim 8 , wherein (i) the subject is a subject pre-diagnosed with IS; (ii) the subject is asymptomatic; and/or (iii) the subject has at least one family member suffering from IS.
10. The composition of claim 8 , wherein the PBMC sample comprises fresh PBMCs.
11. The composition of claim 8 , wherein the ligand is a synthetic ligand.
12. The composition of claim 8 , wherein said ligand for Gi stimulation is selected from the group consisting of: (a) 1-[3-(3,4-Methylenedioxyphenoxy)propyl]-4-phenyl-piperazine (BP554 ) maleate for the 5-HT1A receptor; (b) 5-bromo-N-(4,5-dihydro-IH-imidazol-2-yl)-6-quinoxalinamine (UK14304 ) for the a2-AD receptor; (c) 1-Deoxy-1-[6-[[3-iodophenyl)methyl]amino]-9H-purin-9-yl]-N-methyl-β-D-ribofuranuronamide (IB-MECA) for the A3 receptor; (d) Lysophosphatidic acid (LPA) for the LPA receptor; (e) (2S)-2-[[2-[[(2R)-2-[[(2S)-2-amino-3-(4-hydroxyphenyl)propanoyl]amino]propanoyl]amino]acetyl]-methylamino]-N-(2-hydroxyethyl)-3-phenylpropanamide (DMAGO) for the mu-opioid receptor; (f) 1-(6-Amino-9H-purin-9-yl)-1-deoxy-N-ethyl-β-D-ribofuranuronamide (NECA) for the adenosine receptors; (g) somatostatin for the SSTR receptor; (h) peptide MMK-1 for the FPR2 receptor; (i) Apelin-17 for the APJR receptor; (j) quinpirole for the D2 and D3 receptors; and (k) N-Cyclohexyl-7-chloro-1-[2-(4-morpholinyl)ethyl]quinolin-4(1H)-one-3-carboxamide (CB65 ) for the CB2 receptor.
13. The composition of claim 8 , wherein said ligand for Gi stimulation is melatonin, iodomelatonin or phenylmelatonin.
14. The composition of claim 8 , wherein said ligand for Gi stimulation is somatostatin.
15. The composition of claim 8 , further comprising a reagent for measuring cellular cAMP concentration.
16. A kit for classifying a subject having idiopathic scoliosis (IS) or at risk of developing IS comprising:
(a) recombinant osteopontin (rOPN); and
(b) somatostatin;
wherein said kit optionally further comprises in addition to (a) and (b),
(c) a ligand for Gs stimulation; and/or
(d) one or more antibodies for detecting Giα phosphorylation.
17. A composition for classifying a subject having idiopathic scoliosis (IS) comprising:
(a) an isolated peripheral blood mononuclear cell (PBMC) sample from the subject;
(b) osteopontin (OPN); and
(c) a ligand for Gi stimulation;
wherein said composition optionally further comprises in addition to (a), (b) and (c),
(d) a ligand for Gs stimulation;
wherein said OPN is recombinant OPN (rOPN) or is present at a concentration of 0.5 μg/ml.
18. The kit of claim 17 , wherein said ligand for Gs stimulation is isoproterenol.