RU2496150C1 - Method for simulating bone crystallisation in coxarthrosis in vitro - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: method involves preparing mineral phases forming the basis of an inorganic matrix of human bone tissue in an artificial medium close to such in the individuals suffering coxarthrosis. That involves preparing the above simulative medium at pH 7.80+0.05. The medium has thee following composition: CaCl₂ - 1.1766 g/l, Na₂HPO₄12H₂O - 6.5514 g/l, NaCl - 3.1892 g/l, MgCl₂-6H₂O - 0.4764 g/l, NaHCO₃ - 2.2680 g/l, Na₂SO₄ - 1.6188 g/l, KCl - 0.3427 g/l. After 30 minutes, the medium is examined for hydroxylapatite compositionally similar to apatite of the bone intercellular substance in the patients suffering coxarthrosis.

EFFECT: simulating bone mineralisation in coxarthrosis in vitro and detection of the environment promoting mineral phase deposition in the human joint synovial fluid, enables developing the more effective methods to prevent the development of the given disease.

3 tbl, 4 dwg

Description

The field of technology.

The invention relates to the field of experimental medicine, in particular to the field of traumatology and orthopedics, and specifically to methods for modeling pathological processes of formation of mineral phases in bone diseases.

The level of technology.

One of the first places in the severe course and consequences among diseases of the musculoskeletal system of a person is deforming arthrosis of the hip joint (coxarthrosis). It accounts for 4% to 20% of the pathology of motion support, and the frequency of occurrence is from 15 to 17.8 per 10,000 adults, and increases with age (Sidorenko O.A. Socio-hygienic features of the incidence and evaluation of the effectiveness of patients with pathology large joints: Author. Diss. ... Ph.D. Novosibirsk, 2002, 23 pp .; Egorov KS Prediction of the somatic state of patients with coxarthrosis before and after hip replacement: Abstract. Diss. ... Ph.D. St. Petersburg, 2010, 30 p.).

There is no consensus in the literature on the pathogenesis of degenerative-dystrophic changes in coxarthrosis. Researchers in the development of degenerative-dystrophic changes assign a dominant role to different components of the hip joint: cartilage (Astapenko MG, Bayatova VK. The current state of the problem of deforming arthrosis // Rheumatology. 1984. No. 2. P. 61.), Bone tissue (Sharma L. Local Factors in osteoarthritis // Curr Opin Rheumatol, 2001, V.13, No. 5, P.44-446; Shatsillo OI Osteo-cartilaginous autoplasty in the treatment of metatuberculosis coxarthrosis and aseptic necrosis of the femoral head. Diss. MD, St. Petersburg, 1998, 320 pp.), Both cartilage and bone tissue (Luneva S.N. Biokhimiche changes in the tissues of the joints with degenerative-dystrophic diseases and methods of biological correction. Diss ... Dr. Sc. Tyumen: Tyumen State University, 2003, 297 pp.), synovial fluid filling the joint cavity (Matveeva E.L. Biochemical changes in the synovial fluid during the development of degenerative-dystrophic processes in the knee joint: Abstract of thesis ... Biological Sciences Tyumen: Tyumen State. University, 2007, 24 pp.). The issue of the state and formation of the mineral and organic components of the intercellular substance of the bone tissue of the femoral head in this pathology is slightly covered.

To date, to study the pathogenesis of arthrosis, methods for modeling the clinical picture in animals by introducing intraarticular injection have become widespread (patent RU 2117997 C1, IPC G09B 23/28, 11/17/1994; patent RU 2237928 C2, IPC G09B 23/2, 12/16/2002 ; application for invention 2002133907 A, IPC G09B 23/28), creating an external load on the bone in a state of anesthesia (application for invention RU 2010124506, IPC G09B 23/28 (2006.01), 06/15/2010). The value of these methods is not in doubt, however, they are difficult to access for wide clinical practice. The disadvantage of these models is that they are created within the framework of a living animal organism, which does not allow us to study the pathogenesis of coxarthrosis at the molecular level of the structural organization of human bone tissue (Königsberger E., Konigsberger L. Biomineralization - Medical Aspects of Solubility / John Wiley & Sons Ltd The Atrium, Southern Gate, Chichester, West Sussex PO19 8SQ, England, 2006, 312 p. Weiner S., Addadi L., Wagner HD Materials design in biology // Materials Science and Engineering C, 2000, V.11, P .1-8), namely, to study in detail the processes of conversion of the mineral (hydroxylapatite) and organic (collagen matrix) components of the intercellular substance to deleterious matrix essentially fazoobrazovanija for bone mineralization possible phase transitions and chemical transformations taking into account the various states of biological fluids (intercellular and synovial) in contact with bone tissue in coxarthrosis. The complexity of most real physicochemical processes does not solve the described problems in vivo. Their study becomes possible in an artificially created model system close to the physiological environment.

A known method of modeling the process of formation of tartar, formed as a result of precipitation from saliva of phosphates and carbonates of calcium and magnesium into the organic matrix of tartar, which is the core of the formation. Its essence is to grow tartar by placing a healthy tooth in an artificially created model environment similar in composition to the oral fluid of people with dental deposits (patent RU 2007113160 A, IPC G09B 23/28 (2006.01), 04/09/2007). The advantages of this method include the possibility of studying pathological processes in the human oral cavity. This allows you to identify the causes of the development and course of the disease, therefore, to develop effective methods for its treatment.

The main disadvantages of this model is that it allows you to study only pathological processes that occur in the human oral cavity, in particular, with the formation of plaque and growth of tartar. The indicated drawbacks are connected with the fact that the mixed saliva acts in this case as the mineral-forming medium, which differs from the joint synovial fluid in a wide range of pH values (5.6-7.6), in the content of all inorganic ions, and also in the presence of chloride and rhodanide ions in the composition (Zabrosaeva L.I., Kozlov N.B.Biochemistry of saliva. Omsk, 1992, 44 pp.).

There is a method for producing biocompatible materials to replace bone tissue in a synthetic fluid (SBF) environment that simulates the composition of human blood plasma (Jalota S., Bhaduri SB, Tas AC Using a synthetic body fluid (SBF) solution of 27 mM HCO ₃ ^- to make bone substitutes more osteointegrative // Materials Science and Engineering, V.28, 2008, P.129-140). According to this method, the synthesis of biomaterials was carried out by holding collagen sponges (type I collagen) in a solution similar in composition and inorganic ion content to human plasma at physiological temperatures of 37 ° C and pH = 7.4 for 7 days. In this case, the stabilization of the pH of the model SBF solution was carried out using a buffer mixture of Tris-hydroxymethyl-aminomethane and hydrochloric acid (Tris-HCl), which also made it possible to conduct an experiment at a constant concentration of one of the important plasma ions HCO ₃ ^- (27 mmol / L). The method allows to obtain collagen-apatite-calcium phosphate (collagen-Ap-Cap) composites with nanosized particles and a high surface area, which helps to improve osteogenesis when replacing bone defects in a living organism. Its disadvantages include the lack of the ability to study various pathological processes that occur in bone tissue with bone diseases. This method did not associate their course, in particular, bone mineralization during coxarthrosis with a change in the composition of the biological fluid (for example, the effect of the concentration of sulfate ions), and also did not simulate the biological system in bone pathologies.

The objective of the invention is to develop a method for experimental modeling of pathological bone mineralization in joints with coxarthrosis, identifying conditions conducive to the deposition of mineral phases (different in composition from bone tissue) in the joint synovial fluid in order to develop rational preventive, diagnostic and therapeutic measures prevention and development of this disease.

The specified technical result is achieved by the fact that in the method of modeling bone mineralization in coxarthrosis in vitro, using literature data on the electrolyte composition and pH of synovial fluid in people with coxarthrosis and people who have no bone pathologies (control group) (Matveeva E.L. Biochemical changes in the synovial fluid during the development of degenerative-dystrophic processes in the knee joint: Abstract of Diss. ... Dr. Tyumen: Tyumen State University, 2007, 24 pp .; Kirsanov A.I., Dolgodvorov A.F. Leontiev V.G., Gore Bacheva I.A., Romanova V.D. Velichko L.S., Aleksandrov V.V. Concentration of chemical elements in different biological media // Clinical Laboratory Diagnostics, 2001, No. 3, S.16-20), conduct thermodynamic assessment of the probability of formation of sparingly soluble compounds; the deposition of which is hypothetically possible in this biological medium at minimum, average and maximum ion concentrations in a wide range close to physiological pH = 7.00-8.00, establish significant differences in the composition and sequence of the formed mineral phases from synovial fluid in people with coxarthrosis and persons of the control group, identify significant parameters that contribute to the flow of mineralization in the synovial fluid in people with this disease, based on the data obtained, t modeling environment and observed for a long time. Experimental modeling is performed at pH values and ionic strength of the solution corresponding to the synovial fluid of a group of healthy people (pH = 7.40 ± 0.05), and alkaline deviations in patients with coxarthrosis (pH = 7.60 ± 0.05 and pH = 7.80 ± 0.05). Model media also differ from each other in the content of precipitating calcium ions and inorganic phosphorus. Three model media of the following composition are created: 1. CaCl ₂ - 1.3431 g / l, Na ₂ HPO ₄ · 12H ₂ O - 7.4822 g / l, MgCl ₂ · 6H ₂ O - 0.4764 g / l, NaHCO ₃ - 2.2680 g / l , Na ₂ SO ₄ - 1.6188 g / l, KCl - 0.3427 g / l, NaCl - 2.8798 g / l; 2. CaCl ₂ - 1.2432 g / l, Na ₂ HPO ₄ · 12H ₂ O - 6.9452 g / l, NaCl - 2.8798 g / l, other substances as in model medium 1; 3. CaCl ₂ - 1.1766 g / l, Na ₂ HPO ₄ · 12H ₂ O - 6.5514 g / l, NaCl - 3.1892 g / l, other substances as in model medium 1 and 2. After a month in model systems 1 and 2 is formed poorly crystallized hydroxylapatite with impurities of various inorganic phases, and in model 3, hydroxylapatite is similar in composition to the apatite of the intercellular substance (matrix) of the bone tissue of people with coxarthrosis.

The method is illustrated by illustrations:

figure 1 is a diagram of a model experiment in vitro;

figure 2 - the dependence of the supersaturation indices on the pH of the solution for the average concentration range of the synovial fluid of sparingly soluble compounds: Ca ₁₀ (PO ₄ ) ₆ (OH) ₂ (1); β - Ca ₃ (PO ₄ ) ₂ (2); MgHPO ₄ · 3H ₂ O (3); CaHPO ₄ · 2H ₂ O (4); Ca ₄ H (PO ₄ ) ₃ · 2.5 H ₂ O (5); CaCO ₃ - calcite (6); CaCO ₃ - aragonite (7); α - Ca ₃ (PO ₄ ) ₂ (8);

figure 3 - dependence of the change in Gibbs energy of crystallization from the pH of the solution for the average concentration range of the synovial fluid of sparingly soluble compounds: Ca ₁₀ (PO ₄ ) ₆ (OH) ₂ (1); β - Ca ₃ (PO ₄ ) ₂ (2); MgHPO ₄ · 3H ₂ O (3); CaHPO ₄ · 2H ₂ O (4); Ca ₄ N (PO ₄ ) ₃ · 2.5 H ₂ O (5); CaCO ₃ - calcite (6); CaCO ₃ - aragonite (7); α - Ca ₃ (PO ₄ ) ₂ (8);

figure 4 - diffraction patterns of samples synthesized at pH = 7.4 (1); 7.6 (2); 7.8 (3).

The method is as follows.

To assess the possibility of formation of a sparingly soluble compound from synovial fluid prototypes, thermodynamic calculations of supersaturation indices (SI) were performed [Mullin JW Crystallization. Butterworth-Heinemann. Oxford, 1993, P.118-122] and changes in Gibbs crystallization energies (ΔG) [Koutsopoulos S. Hydroxyapatite crystallization in the presence of serine, tyrosine and hydroxyproline amino acids with polar side groups / S. Koutsopoulos, E. Dalas // Journal of Crystal Growth, 2000, No. 216, P.443-449] for sparingly soluble substances, the formation of which is hypothetically possible in model systems Ca ₁₀ (PO ₄ ) ₆ (OH) ₂ (hydroxylapatite), β - Ca ₃ (PO ₄ ) ₂ (vitlocite), Ca ₄ N (PO ₄ ) ₃ · 2.5H ₂ O (octalcium phosphate), α - Ca ₃ (PO ₄ ) ₂ , MgHPO ₄ · 3H ₂ O (nuberite), MgCO ₃ · 3H ₂ O, Mg ₃ (PO ₄ ) ₂ , CaSO ₄ , CaHCO ₄ · 2H ₂ O (brushite), CaCO ₃ (calcite), CaCO ₃ (aragonite), Ca (H ₂ PO ₄ ) ₂ · N ₂ O and Ca (H ₂ PO ₄ ) ₂ .

According to the results of the calculations, it was found that in the pH range under study in the synovial fluid prototypes the formation of the following compounds is possible thermodynamically (Table 1): Ca ₁₀ (PO ₄ ) ₆ (OH) ₂ , β - Ca ₃ (PO ₄ ) ₂ , MgHPO ₄ 3H ₂ O, Ca ₄ N (PO ₄ ) ₃ · 2.5H ₂ O, CaNRO ₄ · 2H ₂ O, CaCO ₃ - calcite, CaCO ₃ - aragonite, α - Ca ₃ (PO ₄ ) ₂ . Moreover, for each system, the pH of the onset of solid phase deposition is different.

Table 1 Ranges of pH values, supersaturation indices and changes in the Gibbs crystallization energy (kJ / mol) of solid phase formation in model solutions at various concentrations of inorganic ions, mmol / l Minimum Medium Maximum No. Compound concentration concentration concentration pH IS -ΔG pH IS -ΔG pH IS -ΔG one Ca ₁₀ (PO ₄ ) ₆ (OH) ₂ 7.00-8.00 0.80-1.36 4.17-7.11 7.00-8.00 0.89-1.45 4.66-7.60 7.00 8.00 0.95-1.51 4.93-7.90 2 β-Ca ₃ (PO ₄ ) ₂ 7.00-8.00 0.21-0.75 1.07-3.90 7.00-8.00 0.32-0.86 1.65-4.48 7.00 8.00 0.39-0.93 2.01 -4.84 3 Ca ₄ N (PO ₄ ) ₃ · 2.5H ₂ O 7.10-8.00 0.01-0.34 0.02-1.8 7.00-8.00 0.06-0.45 0.33-2.33 7.00 8.00 0.13-0.51 0.65 -2.65 MgHPO ₄ -3H ₂ O 7.00-8.00 0.07-0.24 0.36-1.28 7.00-8.00 0.18-0.36 0.95-1.88 7.00 -8.00 0.25 -0.42 1.3-2.22 four CaHPO ₄ 2H ₂ O 7.15-8.00 0.01-0.14 0.01-7.10 7.00-8.00 0.10-0.28 0.52-1.44 7.00 -8.00 0.19-0.36 0.97-1.90 5 CaCO ₃ (calcite) 7.33-8.00 0.01-0.36 0.02-1.88 7.26-8.00 0.01-0.40 0.006 -2.07 7.21 -8.00 0.01 -0.43 0.02-2.23 6 CaCO ₃ (aragonite) 7.45-8.00 7.45-0.30 0.02-1.53 7.38-8.00 0.01-
0.33 0.01-1.73 7.33 -8.00 0.01-0.36 0.02-1.89 7 α-Ca ₃ (PO ₄ ) ₂ - - - 7.89-8.00 0.01-0.06 0.01-0.30 7.75 -8.00 0.01-0.13 0.02-6.60 8

For a visual comparison of supersaturation indices and changes in Gibbs crystallization energies for three model systems at corresponding pH values of 7.4; pH = 7.6 and pH = 7.8 within the framework of a single coordinate space, graphical functional dependencies of the form SI = ƒ (pH) and ΔG = ƒ (pH) were constructed (Figs. 2, 3). It has been established that the greatest thermodynamic probability of synovial fluid formation in prototypes is characteristic of hydroxylapatite, which is the main component of the intercellular substance of human bone tissue. The values of the thermodynamic parameters (SI and ΔG) of this poorly soluble phase differ from others by 1.6 or more times, which manifests itself to a greater extent at a higher pH = 7.8 (Table 1).

From the data of FIG. 2 and FIG. 3, it follows that under model conditions, the vitlokite β-Ca ₃ (PO ₄ ) ₂ is a stable phase, which in bone mineralization acts as a precursor phase for the formation of hydroxylapatite. In this case, the precipitation of α-Ca ₃ (PO ₄ ) ₂ and calcium carbonates is unlikely, the remaining compounds are intermediate stable and the degree of their participation in the formation of the inorganic component of the intercellular matrix of bone tissue depends on the concentration of precipitating ions, ionic strength, acidity of the simulated medium.

At pH = 7.40 ± 0.05 of the model solution, the thermodynamic probability of formation decreases in the order MgHPO ₄ · 3H ₂ O> Ca ₄ N (PO ₄ ) ₃ · 2.5H ₂ O> CaHPO ₄ -2H ₂ O and is similar for the whole concentration range (table 2). This indicates that at a given pH of the medium, the influence of ionic strength can be neglected.

table 2 Values of supersaturation indices of sparingly soluble compounds of model solutions at various pH values and minimum (1), average (2), maximum (3) inorganic ion concentrations, mmol / l No. pH 7.4 7.6 7.8 Concentrations ^∗ one 2 3 one 2 3 one 2 3 one Ca ₁₀ (PO ₄ ) ₆ (OH) ₂ 1,04 1.14 1,2 1.15 1.25 1.31 1.26 1.35 1.41 2 β-Ca ₃ (PO ₄ ) ₂ 0.45 0.56 0.63 0.55 0.66 0.73 0.65 0.76 0.83 3 Ca ₄ N (PO ₄ ) ₃ · 2.5 H ₂ O 0.14 0.24 0.3 0.21 0.31 0.38 0.28 0.38 0.44 four MgHPO ₄ · 3H ₂ O 0.17 0.28 0.35 0.2 0.32 0.38 0.23 0.34 0.41 5 CaHPO ₄ · 2H ₂ O 0.06 0.2 0.29 0.09 0.23 0.32 0.12 0.26 0.35 6 CaCO ₃ (calcite) 0.05 0.08 0.11 0.15 0.19 0.22 0.26 0.29 0.32 7 CaCO ₃ (aragonite) -0.02 0.02 0.05 0.09 0.12 0.16 0.19 0.23 0.26 8 α-Ca ₃ (PO ₄ ) ₂ -0.35 -0.24 -0.17 -0.25 -0.14 -0.07 -0.15 -0.04 0,03

With an increase in the pH of solutions, the phases with the highest hydrogen density become the most thermodynamically stable. So, at minimum ion concentrations pH = 7.60 ± 0.05, the phase preceding the precipitation of nyuberite is octacalcium phosphate, and nuberite is brushite: Ca ₄ N (PO ₄ ) ₃ · 2.5H ₂ O> MgHPO ₄ · 3H ₂ O> CaHPO ₄ · 2H ₂ O. At pH = 7.80 ± 0.05, this pattern can be traced more clearly.

With increasing ion concentrations in both cases, the forces of interionic repulsion increase, which leads to a decrease in activity coefficients and an increase in the probability of the formation of the most soluble and thermodynamically unstable phase. Following the precipitation of Ca ₄ H (PO ₄ ) ₃ · 2.5H ₂ O, successive formation of compounds is noted, the solubility of which decreases in the order: MgHPO ₄ · 3H ₂ O> CaHPO ₄ · 2H ₂ O (Table 2, FIG. 2).

It can be assumed that the key parameters that can contribute to mineralization in the joint synovial fluid during coxarthrosis and, consequently, the development of this disease are an increase in pH, ionic strength and concentration of precipitating ions. In this case, optimal conditions are created for the formation of hydroxylapatite, the main mineral component of the intercellular matrix of human bone tissues.

Thus, factors are established that contribute to the deviation of the synovial fluid parameters from the norm and the formation of mineral phases in the bone tissue of the joints with the participation of this biological medium.

Then, to confirm the results of thermodynamic calculation, a model experiment was carried out. Model systems were synovial fluid prototypes with an average concentration of inorganic ions at three pH values corresponding to a group of healthy people (pH = 7.4) and patients with coxarthrosis (pH = 7.6 and pH = 7.8).

For the preparation of model solutions, salts of the grade of analytical grade and chemical grade were used. and distilled water. Salts and their amount were selected so that the concentration of their ions in solution and ionic strength were as close as possible to these parameters of the simulated system, namely, the synovial fluid. In this case, the obtained ionic strength differed from the initial value by no more than 0.03. For each series of experiments, solutions were prepared containing cations and anions, with the joint presence of which under these conditions poorly soluble compounds are formed (working solution 1 and 2, Fig. 1). After mixing the equivalent volumes of solutions in a volumetric flask, the achievement of the specified values of the ionic strength was adjusted by adding a certain mass of a sample of crystalline NaCl and adjusted to the mark with distilled water. The resulting solution with pH = 7.50 ± 0.05 was poured into a conical flask with a ground stopper and the pH was adjusted to the required physiological values by adding 20% NaOH solution or concentrated HCl. In order to prevent a decrease in the concentration of carbonate ions in the solution (due to hydrolysis in an acidic medium), the required amount of NaHCO _{3 was} added at pH 5.5-6.0. Flasks with ready-made solutions of the composition: 1. CaCl ₂ - 1.3431 g / l, Na ₂ HPO ₄ 12H ₂ O - 7.4822 g / l, MgCl ₂ 6H ₂ O - 0.4764 g / l, NaHCO ₃ - 2.2680 g / l, Na ₂ SO ₄ - 1.6188 g / l, KCl - 0.3427 g / l, NaCl - 2.8798 g / l; 2. CaCl ₂ - 1.2432 g / l, Na ₂ HPO ₄ · 12H ₂ O - 6.9452 g / l, NaCl - 2.8798 g / l, other substances as in model medium 1; 3. CaCl ₂ -1.1766 g / l. Na ₂ HPO ₄ · 12H ₂ O - 6.5514 g / l, NaCl - 3.1892 g / l, other substances, as in model medium 1 and 2.

Covered with stoppers and left to crystallize at room temperature for 30 days.

To determine the phase and chemical composition of the obtained solid phases, X-ray phase analysis (XRD), IR spectroscopy, and chemical analysis methods were used. The quantitative determination of cations and anions in the solid phase was carried out by the method of residual concentrations by the difference between the initial and final concentrations of precipitating ions in the model solution, where the content of Ca ²⁺ , Mg ²⁺ and inorganic phosphorus ions was analyzed. The concentrations of Ca ²⁺ and Mg ²⁺ were determined according to RD 52.24.403-2007 [RD 52.24.403-2007. Mass concentration of calcium in water. The measurement procedure by the titrimetric method with Trilon B], and inorganic phosphorus based on GOST 18309-72 [GOST 18309-72. Drinking water. Method for determination of polyphosphate content].

Analysis of the solid phases obtained during the simulation at medium electrolyte concentrations and physiological pH values of the synovial fluid confirms the data of thermodynamic calculations. On x-ray samples (figure 4), the main mineral component is a thermodynamically stable phase - hydroxylapatite.

замещающих тетраэдры $$P{O}_4^{3-}$$

в структуре апатита кости [Ньюман У., Ньюман М. Минеральный обмен кости / Перевод с англ. О.Я. Терещенко, Л.Т. Туточкиной. Под ред. Н.Н.Демина. М.: Иностранная литература, 1961, 270 с.], что возможно приводит к снижению кристалличности гидроксилапатита при коксартрозе и отрицательно сказывается на физико-химических свойствах костной ткани головки бедренной кости.At pH = 7.4, the presence of poorly soluble impurity compounds is noted: Ca ₄ N (PO ₄ ) ₃ · 2.5H ₂ O, MgHPO ₄ · 3H ₂ O, CaHPO ₄ · 2H ₂ O, β - Ca ₃ (PO ₄ ) ₂ . With increasing pH, their content decreases and, at pH = 7.8 ± 0.05, the crystalline component of precipitation is hydroxylapatite. Therefore, it can be assumed that in addition to β - Ca ₃ (PO ₄ ) ₂ , which is involved in bone mineralization by phases precursor to the formation of hydroxylapatite during coxarthrosis, calcium phosphates such as octalcium phosphate and brushite can act. Calcium carbonates are absent in the diffraction patterns, which may be caused by small supersaturation indices of these compounds and low thermodynamic stability. These carbonates can act as a source of ionized ions. $$\mathrm{<figure-callout\; id="3"\; label="C\; O"\; filenames="00000007.png"\; state="\{\{state\}\}">C\; </figure-callout>}{O}_{}^{}{}_3^{2-}$$

substituting tetrahedra $$P{O}_{\mathrm{four}}^{3-}$$

in the structure of bone apatite [Newman W., Newman M. Mineral bone metabolism / Translation from English. O.Ya. Tereshchenko, L.T. Tutochkina. Ed. N.N.Demina. M .: Foreign literature, 1961, 270 pp.], Which possibly leads to a decrease in the crystallinity of hydroxylapatite in coxarthrosis and adversely affects the physicochemical properties of the bone tissue of the femoral head.

Processing of chemical analysis data showed (Table 3) that the precipitate closest to stoichiometric (Ca / P = 1.67) is crystallization at pH = 7.80 ± 0.05. This pH value corresponds to the extreme value of the acidity of the synovial fluid during coxarthrosis.

Table 3 Residual concentrations and composition of solid phases in modeling average concentrations of inorganic ions and physiological pH values of human synovial fluid No. pH Source ratios Determined in solution, mmol / l Ratios for solid phases Ca / P Mg / Ca Ca ²⁺ Mg ²⁺

$$P{O}_{\mathrm{four}}^{3-}$$

Ca / P Mg / Ca one 7.40 1.75 0.09 0.70 0.15 2,53 1.88 0.08 2 7.60 1.75 0.10 0.50 0.50 2.15 1.87 0.06 3 7.80 1.75 0.10 0.50 1.00 1,57 1.82 0.01

The Ca / P coefficient of the obtained compound is minimal, and Mg / Ca is 6-8 times lower than the others. This indicates that almost all magnesium ions remained in solution and did not participate in the formation of a solid phase. In this case, the atomic ratio of the Ca and P sediment corresponds to the value of the Ca / P coefficient obtained by us in the study of bone tissues of people with cocartrosis, i.e. the proposed model system allows reproducing the conditions under which mineralization occurs with the participation of human synovial fluid.

Thus, the inventive method allows in model conditions to identify parameters that, when deviated from the norm, lead to mineralization in the bone tissue of the joint with synovial fluid, and create a model system with which you can study the effectiveness of the effects of drugs for the prevention and treatment of coxarthrosis.

Claims (1)

A method for modeling bone mineralization processes that occur during in vitro coxarthrosis, including the production of mineral phases that form the basis of the inorganic matrix of human bone tissue in an artificially created environment similar to that of people with coxarthrosis, while preparing the specified model medium at a pH of 7.80+ 0.05 and the medium has the following composition: CaCl ₂ - 1.1766 g / l, Na ₂ HPO ₄ · 12H ₂ O - 6.5514 g / l, NaCl - 3.1892 g / l, MgCl ₂ · H ₂ O - 0.4764 g / l, NaHCO ₃ - 2.2680 g / l, Na ₂ SO ₄ - 1.6188 g / l, KCl - 0.3427 g / l; after 30 days, hydroxylapatite is determined in this medium, which is close in composition to the apatite of the intercellular substance of the bone tissue of patients with coxarthrosis.

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