Laboratory markers of bone metabolism in different conditions of adaptive reserves of the body and disorders of the musculoskeletal system

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BACKGROUND

The profession of a firefighter is characterized by a clear risk of reducing the functional reserves of the body. Specialists whose activities are associated with the effects of physical and chemical factors and the need to be in constant readiness for immediate action regularly and for a long time experience overexertion. At the same time, decreased concentration associated with day and night shifts is a common cause of injury. The results of the screening assessment of the state of the musculoskeletal system among employees of the Federal Fire Service of the State Fire Service (FPS GPS) EMERCOM of Russia from among the operational staff of the special fire department showed that more than 20% of the surveyed have various complaints of disorders of the musculoskeletal system and need a comprehensive assessment of the condition of the musculoskeletal system, taking into account the general physiological and psychophysiological status.

Laboratory markers of bone metabolism play a significant role in the early diagnosis and prevention of diseases of the musculoskeletal system as participants in the overall adaptive response of the body to stress factors of various nature. It is known that bone tissue, regardless of age, is constantly undergoing two mechanisms of restructuring: modeling and remodeling. Modeling determines the microstructure of the bone during its growth. Remodeling consists of resorption of local areas and filling of the formed defects with newly formed bone tissue.  A specific marker of bone resorption is deoxypyridinoline in urine, a pyridine derivative that serves as a "molecular crosslinking" that stabilizes the structure of collagen fibrils. The release of pyridine derivatives from bone tissue is a reliable indicator of the breakdown of mature collagen. The resorption phase lasts up to 12 days and ends with the death of osteoclasts, followed by the involvement of osteogenic cells and their differentiation into osteoblasts. Osteoblasts synthesize a new bone matrix and participate in its calcification, and then, as the intercellular substance accumulates and orientates spatially, they become osteocytes. The full cycle of bone remodeling lasts 3-6 months, with osteosynthesis prevailing over resorption. Remodeling allows you to adapt the volume, shape and density of bone to the current loads, adjust and update the microarchitectonics of the tissue. This process is also part of the system that ensures the maintenance of an optimal concentration of minerals in body fluids.

A specific marker of osteosynthesis is osteocalcin, a calcium-binding non-collagen protein of the intercellular matrix, which is produced by mature osteoblasts. Osteocalcin makes up up to a quarter of the non-collagen protein matrix of bone and is represented by two forms: carboxylated and bioactive subcarboxylated. Osteocalcin synthesis by osteoblasts is stimulated by vitamin D, and the concentration of the active form of this protein is regulated with the participation of vitamin K. When interpreting the results of determining the level of osteocalcin in the blood, it is necessary to take into account that this protein performs several functions. First of all, it is an intercellular matrix protein, and its level in the blood reflects the activity of osteoblasts and the intensity of bone formation. However, one tenth of the formed osteocalcin enters the bloodstream directly, which is necessary for its intersystemic adaptive effects, including its effect on energy metabolism and neurotransmitters. [1, 2]. So, it is currently known that osteocalcin has hormonal activity: it increases the proliferation of beta cells of the pancreas and insulin secretion, increases the sensitivity of target organs to insulin, stimulates the production of testosterone and cortisol, affects the conduction of impulses by pyramidal brain cells during memory modulation and reduces anxiety. The effects of osteocalcin in adaptation to physical exertion are realized in an increase in muscle strength due to increased absorption and catabolism of glucose and fatty acids by myofibrils, as well as due to an increase in the number and size of mitochondria [1, 3]. Biological activation of osteocalcin is stimulated during bone resorption, while the concentration of the active form of osteocalcin, as well as markers of resorption, it doubles even after a single physical exertion. During aerobic exercise, osteocalcin levels double at the moment when insulin concentration reaches a minimum [4]. An increase in osteocalcin production in response to stress is a fast-acting physiological adaptive mechanism, and relative to the musculoskeletal system, this hormone acts as a cortisol antagonist. Consequently, the level of osteocalcin in the blood depends on the predominance of certain mechanisms for maintaining physiological balance in the body and, within the limits of reference values, does not serve as a significant marker of the severity of diseases of the musculoskeletal system.

Vitamin D is of great importance in the processes of bone tissue remodeling, which stimulates the production of osteocalcin, regulates calcium-phosphorus metabolism and directly affects bone regeneration. Its free form, 1,25(OH)2D3, participates in the regulation of bone resorption, differentiation of osteogenic cells, mineralization of osteoid and fracture healing [5, 6]. The metabolite of vitamin D – 24R,25(OH)2D – is considered as a "critical catalyst" in stimulating bone recovery after injury [1], since under its influence the signaling molecule lactosylceramide is synthesized, which increases the area of the regenerate. Data from a population-based study have been published, which revealed a high prevalence of vitamin D deficiency (40%) and severe deficiency (11%) in adult patients with fractures [2]. Nevertheless, there are works that do not confirm this relationship [5]. It is also known that vitamin D deficiency is manifested not only by a shift in the physiological balance of bone remodeling processes, but also by fibromyalgic syndrome, myalgia and arthralgia.Таким образом, функции многих маркеров метаболизма костной ткани не ограничиваются опорно-двигательной системой, а играют существенную роль в процессах адаптации организма к стрессовым нагрузкам различной природы: физической, климатической и эмоциональной.

In this case, the bone performs the function of a hormone-producing organ and is a participant in adaptation. An urgent task is to establish laboratory criteria for the depletion of bone-mediated adaptation mechanisms, which may be important for the prevention of clinically pronounced disorders of the musculoskeletal system.

AIM

to identify early markers of adaptation of the musculoskeletal system to the effects of stressful occupational factors in employees of the Federal Fire Service of the State Fire Service (FPS GPS) EMERCOM of Russia.

 

MATERIALS AND METHODS

Characteristics of the groups. 105 FPS GPS employees (men) were under medical supervision, the average age was 34,7±0,4 years. For the laboratory examination, the men were divided into three groups depending on the condition of the musculoskeletal system according to the method of "Assessment of disorders of the musculoskeletal system" [7] and into three groups depending on professional experience (табл. 1). The groups did not differ in age. The phenotypic and physiological characteristics of the groups examined according to the medical examination data are given in Table. 1.

 

Table 1. Criteria for the distribution of the examined into groups and general characteristics of the groups according to the dispensary observation data

 

Group attribute	Disorders of the musculoskeletal system			Length of work experience, years
Group criteria	No	Mild	Severe	≤ 5	6-14	≥ 15
Group, №	1	2	3	4	5	6
Number of people	71	20	14	38	45	22
Body mass index	↑	↑	↑	N	↑	↑
Waist–hip ratio	N	N	↑	N	↑	↑
Stange’s test	N	N	N	N	N	N
Genchi’s test	N	N	N	N	↓	N

"N" – inside the reference range, "↑" – above the reference range, "↓" – below the reference range

 

The assessment of the functional state of the cardiorespiratory and cardiovascular systems was based on breath-holding tests on inspiration (Stange test) and exhalation (Gencha test). The lower limits of the reference range of the sample results were considered to be 40 s and 35 s, respectively. Body mass index (BMI) (kg/m²) was calculated using the formula: height (m)/ [body weight (kg)]² and was estimated in accordance with the reference range of 18.5-25 kg/m² of the World Health Organization (WHO). The waist/hip index was calculated as the ratio of waist (m) to hip (m) and was estimated in accordance with the WHO norm limit for men -0.9.

Materials. The level of all parameters, with the exception of osteocalcin and deoxypyridinoline, was studied in venous blood samples in primary vacuum tubes on the day of blood sampling. The blood serum was separated by centrifugation at 3000 rpm for 10 minutes. To study the osteocalcin level, serum aliquots were taken from primary vacuum tubes into 1.5 ml Eppendorf tubes, which were stored at -20 °C. Defrosting was performed once, and the osteocalcin concentration in all aliquots was determined simultaneously. The concentration of deoxypyridinoline was determined in the second portion of morning urine taken into vacuum containers on the day of blood sampling. Urine samples were centrifuged at 3000 rpm for 10 minutes.

 

Methods of the laboratory research. The list of the studied indicators, the analytical systems used and the reference values are presented in Table 2.

 

Table 2. The list of studied indicators, the used analytical systems and reference values

 

Parameter, units of measurement	The reference range for men	Analytical system
Parathyroid hormone, pmol/l	1,2–7,6	Immulite XPi, Siemens, США
Dehydroepiandrosterone sulfate, mmol/l	2,2–15,2
Calcitonin, pg/ml	< 18,2
Deoxypyridinoline, nmol/mmol creatinine	2,3–5,4
Cortisol, nmol/l	185,0–624,0	Access 2, США Beckman Coulter
25-OH-vitamin D, nmol/l	50,0–75,0 –   insufficient 75,0–250,0 –  sufficient
Osteocalcin, ng/ml	9,6–40,8	IDS N-MID Osteocalcin

 

 

Previously, we found [8] that the ratio of concentrations of dehydroepiandrosterone sulfate (DHEAS) and cortisol serves as an objective assessment of the state of the body's adaptive reserves. The value of the DHEAS/cortisol index was interpreted as follows: <1.1 – adaptive reserves are depleted; from 1.1 to 2.1 – adaptive reserves are consumed; >2.1– adaptive reserves are preserved.  The insulin resistance index (HOMA-IR) was calculated using the formula: [insulin (µme/ml)×glucose (mmol/L)]/22.5.

We also calculated the ratio of osteocalcin and insulin concentrations (osteocalcin/insulin index), guided by modern concepts of osteocalcin as a hormone with an anabolic effect that increases the sensitivity of target organs to glucose, the peak concentration of which in the blood in physiological equilibrium falls at a minimum concentration of insulin.

Statistical methods. Statistical analysis was performed using the program "Statistica 10.0" (license AXA009K287210FAACD-B). The correspondence of quantitative laboratory results to the normal distribution was assessed using the Shapiro-Wilk criterion. When describing the data obtained, the arithmetic mean (M) and the standard error of the arithmetic mean (m) were indicated. The significance of the differences in paired comparisons was assessed using the Mann–Whitney U-test. The critical significance level (p) was assumed to be 0.05 when testing statistical hypotheses.

RESULTS

The average values of the studied indicators in all groups were within the reference range.  The vitamin D level was assessed as "insufficient", but it was also within the limits of the population norm.

The main results of the laboratory examination of the FPS GPS employees with various conditions of the musculoskeletal system are presented in Table 3.

 

Table 3. The main results of the laboratory examination of FPS GPS employees with various conditions of the musculoskeletal system

 

Parameter, units of measurement	Group, №			Level of significance of the differences between groups 1,2; 1,3; 2,3;
	1	2	3
Osteocalcin, ng/ml	17,21 ± 1,33	10,89 ± 0,56	17,93 ± 2,85	0,004; 0,480; 0,067;
Osteocalcin/Insulin, index	6,09 ± 0,81	3,0 ± 0,38	3,27 ± 1,1	0,023; 0,058; 0,323;
Insulin, mME/l	5,02 ± 0,55	5,18 ± 1,07	10,2 ± 2,93	0,111; 0,007; 0,127;
НОМА-IR, index	1,06 ± 0,12	1,45 ± 0,3	2,15 ± 0,62	0,086; 0,007; 0,199;
25-OH-vitamin D, nmol/l	60,8 ± 1,86	53,36 ±1,82	65,1 ± 4,22	0,062; 0,568; 0,049;

 

Groups 1,2,3 did not differ in the following indicators: BMI, parathyroid hormone, calcitonin, DHEAS/Cortisol and deoxypyridinoline. A decrease in osteocalcin levels (10.89 ± 0.56 ng/ml) was detected in individuals with moderate musculoskeletal system disorders compared with its average level (17.21 ± 1.33 ng/ml) in group 1. However, in group 3, the tendency to decrease this marker of osteosynthesis did not persist, which is probably due to the activation of bone formation processes in individuals with severe pathology of the musculoskeletal system.

The main results of the laboratory examination of the FPS GPS employees with various professional experience are presented in Table 4.

 

Table 4. The main results of the laboratory examination of FPS GPS employees with various length of work experience

 

Parameter, units of measurement	Group, №			Level of significance of the differences between groups 4,5; 4,6; 5,6;
	4	5	6
Body mass index, kg/m2	24,76 ± 0,35	26,97 ± 0,52	28,33 ± 0,81	0,001; 0,001; 0,239
Osteocalcin, ng/ml	18,5 ± 2,15	15,26 ± 1,16	13,1 ± 1,58	0,386; 0,049; 0,133;
Osteocalcin/Insulin, index	6,47 ± 1,03	4,28 ± 0,59	3,83 ± 0,8	0,049; 0,031; 0,369;
Calcitonin, pg/ml	2,52 ± 0,21	3,29 ± 0,26	4,44 ± 0,52	0,001; 0,001; 0,001

 

Groups 4,5,6 did not differ in the following indicators: parathyroid hormone, calcitonin, insulin, HOMA-IR, DHEAS/Cortisol, vitamin D and deoxypyridinoline. In individuals with short experience (group 4), the average osteocalcin level (18.5 ± 2.15 ng/ml) was comparable to the median of the reference range (19.8 ng/ml) and differed from the lower concentration of this protein in individuals in group 6.  A decrease in the osteocalcin/insulin index, in contrast to the dynamics of insulin concentration, was revealed not only when comparing groups 4 and 6, but also groups 4 and 5. However, the correctness of the assessment of these results is compromised by significant differences in BMI between the groups.  Similar changes in the level of the osteocalcin/insulin index were found when comparing groups 1 and 2 (Table 3).

Osteocalcin concentrations were significantly higher in individuals with active expenditure of adaptive reserves compared to those examined whose adaptation reserves were preserved (Table 5).

 

Table 5. The level of osteocalcin in the blood serum of FPS GPS employees with a different state of adaptive reserves of the body

 

Parameter	DHEA-S/Cortisol, index		Level of significance of the differences
	< 2,1	>2,1
Number of people	38	67	-
Age, years	32,8 ± 0,6	32,7 ± 0,8	0,594
Osteocalcin, ng/ml	20,15 ± 2,18	13,21 ± 0,66	0,002

 

Exceeding the reference limits of osteocalcin levels was also detected only in those examined with a DHEAS/cortisol index <2.1, among whom it accounted for 8% of cases.

Since the reason for the contradictory dynamics of osteocalcin levels in groups 1-3 could be the small number of the third group relative to group 1, we combined individuals with moderate and severe disorders of the musculoskeletal system (Table 6).

 

Table 6. The main results of the laboratory examination of FPS GPS employees with and without disorders of the musculoskeletal system

 

Parameter	Disorders of the musculoskeletal system		Level of significance of the differences
	No	Yes
Number of people	71	34	-
Group, №	1	2,3	-
Body mass index, kg/m2	27,74 ± 1,58	26,87 ± 0,54	0,751
Osteocalcin/Insulin, index	6,09 ± 0,81	3,31 ± 0,51	0,015
Insulin, mME/l	5,02 ± 0,56	7,65 ± 1,33	0,026
НОМА-IR, index	1,06 ± 0,12	1,72 ± 0,31	0,007

 

 

During the comparison, the tendency to increase the levels of insulin and HOMA-IR against a stable BMI was confirmed, and no differences were found in calcitonin and osteocalcin levels. At the same time, there was a distinct decrease in the osteocalcin/insulin index in people with musculoskeletal disorders.

Discussion

The concentrations of deoxypyridinoline and parathyroid hormone did not differ in all six groups, which indicates the absence of specific pathological changes in bone resorption and calcium metabolism. The increase in calcitonin levels is noteworthy and requires further study as professional experience increases (Table 4). This fact can be considered as a compensatory mechanism of osteoblast stimulation against the background of a decrease in osteocalcin, since possible physiological fluctuations in blood calcium levels do not significantly affect calcitonin secretion. 

According to phenotypic characteristics, groups 3 and 6 were characterized by abdominal obesity (Table.1), which may indicate a violation of carbohydrate metabolism in the FPS GPS employees with severe disorders of the musculoskeletal system and extensive professional experience. However, a laboratory examination in group 4 revealed a lower concentration of insulin and a higher sensitivity to insulin, which is characterized by HOMA-IR (Table 4). Groups 1 and 3 did not differ in these indicators (Table 3). In general, the following feature should be noted. If, with increasing professional experience, an increase in BMI was not accompanied by an increase in HOMA-IR and insulin concentration, then as the condition of the musculoskeletal system worsened, the observed increase in HOMA-IR and insulin levels was not accompanied by an increase in BMI. Taking into account the fact that all the detected fluctuations of markers are within the reference ranges, it can be assumed that this feature also corresponds to the physiological limits of adaptation, rather than pathological changes in carbohydrate metabolism.

The change in osteocalcin levels, which characterizes the difference between healthy individuals and those with musculoskeletal system pathology, is consistent with the data of T.T. Tsoriev et al. [9], but needs to be clarified taking into account a statistically insignificant upward trend in its level in group 3. On the other hand, the results of a clinical and laboratory examination of the FPS GPS employees are consistent with modern ideas about the involvement of osteocalcin in enhancing anabolic processes during the stress phase of the adaptive response. It is possible that the physiological enhancement of osteocalcin secretion into the systemic bloodstream is a mechanism that ensures the successful adaptation of the FPS GPS employees to chronic occupational stress. We observed a similar trend of changes in osteocalcin concentration during the dispensary examination of professional football athletes in 2009-2011.  On the contrary, a decrease in osteocalcin concentration within the reference range can be considered as a laboratory sign of depletion of adaptation reserves. Our results are also consistent with those of Pittas et al., who established an inversely proportional relationship between osteocalcin levels and biochemical markers of metabolic syndrome and obesity in men over 65 years of age [10].  So, in our study, there was a clear trend towards multidirectional dynamics of osteocalcin and insulin concentrations, HOMA-IR both with increasing seniority and with the development of disorders of the musculoskeletal system among the staff of the FPS GPS employees.

It is acceptable to assume that the revealed changes in osteocalcin levels are due to a decrease in the intensity of the physiological mechanisms of osteocalcin-mediated adaptation. It is important that the DHEAS/cortisol index exceeded 1.1 in all groups; there was no depletion of adaptive reserves. Thus, the laboratory assessment of the body's adaptive reserves characterizes the examined individuals as resistant to stress, which coincides with the results of functional tests evaluating the reserves of the cardiorespiratory system (Table 2).

 

 

CONCLUSION

The laboratory assessment of the adaptive reserves of the body of the FPS GPS employees coincides with the results of functional tests performed during the dispensary monitoring of their health. The indicators of the state of carbohydrate metabolism do not coincide with its assessment based on phenotypic characteristics, which is probably due to adequate subclinical compensation for obesity due to the preserved reserves of adaptation in young men.

The results of a complex laboratory examination of the FPS GPS employees indicate that they do not have pathological changes in the level of markers of bone metabolism, significant for the diagnosis of clinically pronounced disorders of the musculoskeletal system. Nevertheless, the results confirm the involvement of the musculoskeletal system in polysystem adaptation to stress. Thus, the concentration of osteocalcin within the reference range was higher in individuals with active expenditure of adaptive reserves compared with those examined, whose adaptation reserves were preserved.  There was also a decrease in the osteocalcin/insulin index as the FPS GPS employees increased their professional experience and the condition of the musculoskeletal system worsened.

The data obtained determine the expediency of further studying the role of osteocalcin and the osteocalcin/insulin index in the early diagnosis of musculoskeletal diseases and maintaining tolerance to occupational stress in people in dangerous professions. Markers have the potential to be used in preventive action programs aimed at timely correction of stress-induced adaptive variations in homeostasis.

About the authors

Natalya Aleksandrovna Alkhutova

Всероссийский центр экстренной и радиационной медицины им. А.М. Никифорова МЧС России

Email: nalhutova@yandex.ru
ORCID iD: 0000-0002-6268-8969
SPIN-code: 8732-2680

PhD Biol. Sci., Senior Research Associate Laboratory of Clinical Laboratory Department

Russian Federation, The Nikiforov Russian Center of Emergency and Radiation Medicine, EMERCOM of Russia

Nadezhda Alekseevna Kovyazina

The Nikiforov Russian Center of Emergency and Radiation Medicine, EMERCOM of Russia

Author for correspondence.
Email: nakovzn@gmail.com
ORCID iD: 0000-0002-0482-0802

Dr. of Medicine, Head of the Laboratory of Serological Research and Allergodiagnostics

Russian Federation

Sergey Sergeevich Aleksanin

the Nikiforov Russian Center of Emergency and Radiation Medicine, EMERCOM of Russia

Email: medicine@nrcerm.ru
ORCID iD: 0000-0001-6998-1669
SPIN-code: 1256-5967

Доктор медицинских наук, профессор, член-корреспондент РАН, директор

Russian Federation, Russia, 194044, St. Petersburg, Academica Lebedeva Str., 4/2

Victor Jur’evich Ribnikov

The Nikiforov Russian Center of Emergency and Radiation Medicine, EMERCOM of Russia

Email: rvikirina@mail.ru
ORCID iD: 0000-0001-5527-9342
SPIN-code: 3720-0458

доктор медицинских наук, доктор психологических наук профессор, заместитель директора по научной и учебной работе, медицине катастроф

Russian Federation, Russia, 194044, St. Petersburg, Academica Lebedeva Str., 4/2)

Denis Fyodorovich Magdanov

The Nikiforov Russian Center of Emergency and Radiation Medicine, EMERCOM of Russia

Email: magdanov74@mail.ru
ORCID iD: 0009-0004-5403-8888
SPIN-code: 3922-6243

Заведующий отделением ортопедии отдела травматологии и ортопедии

Russian Federation, Russia, 194044, St. Petersburg, Academica Lebedeva Str., 4/2)

References

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