K Haranadh¹, Bachu Rajendra Prasad^2*

¹Associate Professor, ²Assistant Professor, Department of Orthopaedics, Kamineni Institute of Medical Sciences, Narketpally, Nalgonda (Dt.), Telangana, INDIA.

Email: haranadh112233@gmail.com, raju.kims@gmail.com 

RESULTS

The control group comprised 20 healthy individuals (men) who voluntarily agreed to participate in this study, these volunteers were aged between 20–30 years, and they had no history of any systemic diseases and bone fractures within the last five years. The patients group consisted of 20 individuals (men) aged 20–37 years, suffering from proximal tibial fractures.

Table 1: Estimation of Nitric Oxide (NO) levels (µM) in the control and patient groups during recovery.

Data represent by Mean ± SE, *p < 0.05, significant **p < 0.01, very significant.

The concentration of total NO was higher in the serum of all the patients with fractures before and after the procedure (Table 1). The statistically significant differences were observed in the total NO concentrations in the serum between the patients with the control group at p < 0.05.

Table 2: Estimation of Reactive Nitrogen Intermediates (RNI) levels ((nmol/ml) in the control and patients groups during the recovery period.

Data represent by Mean ± SE, *p < 0.05, significant **p < 0.01, very significant.

There were a significant rise in the Reactive Nitrogen Intermediates (RNI) levels during the 7th day and a peak during the 14th day in the patients group compared to the control group (Table 2). The increase in the patients group was significantly more than the control group. Inpatient group RNI levels were decreased almost to the initial level as on day one by the 28th day.

Table 3: Estimation of Citrulline (CIT) levels ((nmol/ml) in the control and patient groups during the recovery period.

Data represent by Mean ± SE, Significant *p < 0.05, **p < 0.01.

There was a significant elevation in the Citrulline (CIT) levels during the 7th day and a peak during the 14th day in the patients group compared to the control groups (Table 3). These levels were decreased almost to the initial values by day 21.

Table 4: Estimation of Malonyldialdehyde (MDA) levels (µmol/ml) in the control and patient groups during the recovery period.

Data represent by Mean ± SE, Significant *p < 0.05, **p < 0.01.

There were a significant increase in the Malonyldialdehyde (MDA) levels by the 14th day in the patients group (Table 4). By the 21st and 28th days, MDA levels decreased to less than values seen on day 1.

Table 5: Estimation of Ferric Reducing Anti-Oxidant Power Assay (FRAP) levels (µmol/ml) in the control and patient groups during the recovery period.

Data represent by Mean ± SE, Significant *p < 0.05, **p < 0.01.

There were a significant rise in the Ferric Reducing Anti-Oxidant Power Assay (FRAP) levels by the 14th day in the patients group compared to the controls (Table 5). By the 28th day, FRAP values again decreased to a level less than those on day 1.

DISCUSSION

In normal healthy bone and muscles, tissues have a significant amount of active eNOS, producing low NO amounts. The local synthesis of this molecule is sufficient to stimulate both these cell types. However, during inflammation, due to the action of inter-alia, proinflammatory cytokines, the activity of iNOS may be restored along with an intensified synthesis of Nitric Oxide (NO). Elevated NO levels in the intracellular fluid and intercellular space may lead to cell apoptosis, including that of pro-osteoclasts and leukocyte deficiency, which contributes to an inhibited alteration of the cytoskeleton.^1-3 Therefore, high concentrations of total NO were observed in the serum of patients with proximal tibial fractures throughout the study period.^4,5 Radiation, pollution, smoking, alcohol addiction and obesity have adverse effects on the biological systems, and the literature [1-6] has supported some benefits of antioxidants to counter these adverse effects. So much is talked about antioxidants and their role in physiological and pathological conditions that it made us think of their importance in our subset of patients who sustained fractures. This study was designed to evaluate oxidative stress after sustaining a fracture. Corbett et al.¹⁴ have shown that increased activity of eNOS in cortical bone and blood vessels at the beginning stages of fracture (on the first day) as well as an increased stimulation of iNOS in the recovery period (2 weeks after injury) in a rabbit model. A similar type of measurement was also reported by Keskin et al.,¹⁵ who observed higher concentrations of total NO in the serum of patients with an isolated femur fracture. Prasad et al.¹⁶ also demonstrated high NO concentrations in the serum of patients with isolated femur fractures and other fractures in long bones apart from the femur. Gokturk et al.¹⁷ evaluated oxidant status during bone healing in rats by using MDA levels in bone specimens to indicate oxidative stress. They noted statistically significant increases in MDA levels on days 7 and 14 after experimentally fracturing the right tibia of the study rats. They concluded that oxidative stress occurs during the second and third weeks after a fracture. Since the MDA levels were evaluated in the bone specimens, they also postulated that this oxidative stress could potentially cause impaired fracture healing.^18-20 The increased values of RNI, cit and MDA by the 7th day, peaking by the 14th day, and the decline over the next two weeks showed a significant level of oxidative stress during the second- and third-weeks post-fracture. The FRAP values, representing the total endogenous antioxidant status, also showed a similar pattern: i.e., rise during the 7th and 14th days and gradual decline by the end of the fourth week. Although we have no clear explanation for this, we presume that it may attempt to combat the rise in oxidant levels. The effect of antioxidants needs to be seen by fixing the timing of surgery. It is also recommended that fracture cases be studied for a more extended period, perhaps until the union is achieved, to see further variations.

CONCLUSIONS

The duration of fracture healing observed in patients with a proximal tibial fracture at different intervals was found to be related with an elevated NO concentration, as well as changes in the concentrations of MDA, indicating that the intensity of local processes is reflected by the changes in the action of NO in the serum. The observations from the present study suggest the applicability of NO's determination in assessing proximal tibial fracture healing. This preliminary study has attempted to create a platform for further research. We assume that the observations of this study may help in understanding fracture healing better.

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