The present investigation was conducted to study the use of local antibiotic and biodegradable bone cement on healing of diaphyseal long bone compound fractures in dogs. The dogs divided into two groups namely Group I, which received local antibiotic treatment and Group II, control group without local antibiotic therapy. The haematological parameters like Total leucocyte count (TLC), differential leucocyte count (DLC), Erythrocyte sedimentation rate (ESR) and serum biochemical parameters such as alkaline phosphatase and C-reactive protein were estimated pre and postoperatively on 0th day, 7th, 14th, 28th, 45th and 60th post-operative day. Blood parameters were significantly higher in group II dogs compared to group I in the post-operative period throughout the research period. The serum alkaline phosphatase values significantly increased from 0th day to14th day and thereafter decreasing to normal values by 60th postoperative day with more in group I. C-reactive protein (CRP) values were high in group II compared to group I.
Alkaline phosphatase enzyme activity and CRP values were assessed during fracture healing to identify fluctuations, if any in fracture healing after rigid stabilization is till date arguable and are regularly evaluated by many authors (Hegade et al., 2007). Biochemical markers of bone turnover are usually divided into two subclasses: bone resorption and bone formation markers. The bone resorption markers are related to osteoclast resorption of matrix and include tartarate resistant acid phosphatase and degradation products of type I collagen in protein matrix especially hydroxyproline, telopeptides etc. Bone formation markers are osteocalcin and bone specific alkaline phosphatase produced from osteoblasts (Delmas, 1995). Even though clinical examination and radiographic studies gives an idea of progress of fracture union, the literal status of bone resorption and bone formation can be achieved over a short time interval using these biochemical markers (Mukhopadhyay et al., 2011). C-reactive protein is an acute phase reactant, which was produced by the liver and released into the blood within a few hours after tissue injury, the start of an infection, or other cause of inflammation and proved helpful as a marker in risk stratification and as an early indicator for infection (Scherer et al., 2001). The haematological parameters like TLC, neutrophil count were assessed to study the presence of any infection in the post-operative period of fracture stabilization (Umeshwori et al., 2015). The present study was carried out to assess the degree of bone formation, pain elicited and infection rate at fracture site postoperatively using different serum and blood parameters after stabilizing the long bone compound fractures in dogs with local biodegradable bone cement-antibiotic impregnated plates and intramedullary pins.
Materials and Methods
Thirty dogs of highly unstable different diaphyseal long bone compound fractures were selected to study and were divided into two groups. In Group I, 24 cases were stabilized by Locking Compression Plate (LCP) or Intramedullary Pin (IMP) based on the bone involved and fracture configuration. While stabilization, impregnation of the implant surface was done with antiobiotic (gentamicin) and bone cement paste (Hydroxyapatite or Poly-D, L-Lactide) in case of locking compression plating technique for radial and tibial fractures and beads made with antibiotic (gentamicin) and bone cement (Hydroxyapatite or Poly-D,L-Lactide) were used in case of intramedullary pinning technique for humeral and femoral fractures. In Group II, Six dogs were stabilized with LCP and IMP for various fractures without use of any local antibiotic and bone cement treatment as control group which received only parenteral antibiotics post-operatively. Blood samples were collected and serum was separated in all the cases before and after surgery on day 0th day, 7th, 14th, 28th, 45th and 60th in all the cases and estimated serum alkaline phosphatase and C-reactive protein. Serum alkaline phosphatase (IU) was estimated by IFCC kinetic assay method (Young, 1997). C-reactive protein values (mg/dl) were estimated by following latex slide and tube test with kit from Span Diagnostics Ltd (Young, 1997). The ESR was estimated using wintrobe method. The data regarding serum biochemical parameter and haematological values were subjected to standard statistical analysis using one way ANOVA as described by Snedecor and Cochran (1994) using SPSSR 15 software package.
Results and Discussion
In group I, the mean total leucocyte count in entire study period was within normal range except for the count in preoperative period. Neutrophilia was observed in dogs pre and postoperatively showing a decreasing trend by 60th postoperative day. The erythrocyte sedimentation rate increases and reached peak on 7th postoperative day, later reaching normal values by 14th postoperative day. There was mild increase in the mean value of alkaline phosphatase post-operatively which reached peak on 14th postoperative day and later decreased and remained within normal range in all the dogs. The mean C-reactive protein were relatively high preoperatively and showed peak value on 7th postoperative day, when it showed value above normal (Table 1).
Table 1: Haematology and Blood biochemistry levels in group 1 animals
|Alkaline phosphatase (IU)||98.50±0.99||126.83±0.94||138.83±1.25||104.83±1.01||90.33±1.38||84.33±0.88|
|C-reactive protein (μg/ml)||34.17±1.79||43±1.41||24.16±0.79||19.5±0.43||12.50±0.76||8.83±0.60|
In Group II (control group), the values differed significantly compared to Group I at any instance in post-operative day. High Mean leucocyte count with marked neutrophilia and high C-reactive protein values were observed upto 28th post-operative followed by decreasing trend. Alkaline phosphatase reached peak on 14th post-operative day (Table 2).
Table 2: Haematology and Blood biochemistry levels in group 2 animals
|Alkaline phosphatase (IU)||99.45±1.37||108.23±0.67||116.67±2.54||97.79±2.87||85.46±1.55||86.33±0.47|
|C-reactive protein (μg/ml)||33.15±0.85||46.14±1.70||30.33±1.41||26.71±1.53||21.54±0.59||15.23±1.66|
The serum alkaline phosphatase values significantly increased from preoperative day to 14th day and there after the levels decreased reaching normal by 60th day. Increase in the alkaline phosphatase activity was observed during osteoblastic activity with high increase in most of compression methods of internal fixation (Guyton, 1981). Increase in serum alkaline phosphatase level may be due to increased chondroblastic proliferation to cause bone formation during fractured bone repair and also maximum contribution was from the periosteum of destructed bone which was a rich source of serum alkaline phosphatase. The findings were in concurrence with those of Guyton (1981), and Maiti et al. (1999) Ghosh et al. (2003), Tembhurne (2006). Rani et al. (2012) observed an increase in alkaline phosphtase activity upto 21st post-operative day and returned to near base value on 60th post-operative day. Contrary to this, Singh et al. (1976) reported increased serum alkaline phosphatase activity throughout the study period which was attributed to the muscle, skin trauma and early stage of bone repair. Komnenou et al. (2005) explained that the detection of specific biochemical markers of bone formation in serum, such as alkaline phosphatase activity can be clinically useful in evaluating the progress of the healing process and serves as an additional tool in predicting fractures at risk of developing a non-union. Based on the values obtained in group I and group II, it was concluded that the chondroblastic activity was in group I, which was attributed to biodegradable bone cement and less infection at fracture site. CRP values differed significantly in different stages of postoperative intervals. At any instance, CRP values in group II were higher compared to group I, owing to prolonged post-operative inflammation in group II. These elevated values might be because of initial pain due to fractures as opined by Caspi et al. (1984) and surgical trauma according to Yamamoto et al. (1993). Awareness of the natural course of the CRP response after fracture and arthropalsty may be helpful in the diagnosis of early post-traumatic and postoperative complications, especially infections (Yamamoto et al., 1992). The values significantly increased from 0 day to 14th day in all the cases and there after the levels decreased.
Erythrocyte sedimentation rate (ESR) increased in the preoperative and initial post- operative period above the normal value and reached peak on 7th post-operative day thereby showed decreasing trend to base value on 60th post-operative day in both the groups. The increase in ESR was attributed to the initial inflammation and infection as the fractures under study were compound fractures. The increase in ESR due to inflammation and infection was also observed by Khan et al. (2011) and Alam et al. (2005). Obiorah et al. (2017) reported increased ESR in a dog affected with parasitic infection compared to a normal healthy dog. Alam et al. (2005) observed mean ESR value in a group of dogs encountered with closed fractures, which was less compared to the mean ESR value in the present study owing to the compound nature of the fractures. The ESR in group I dogs differed significantly from that of control group in post-operative period. Significant neutrophilia with increased total leucocyte count was observed preoperatively in all the dogs, which could be attributed to the stress associated with the fracture leading to muscle damage, hematoma formation or infection at the fracture site in accordance to the findings of Umeshwori et al. (2015) who also reported mild neutrophilia in fracture cases. Sastry (1989) opined that neutrophilia occurred in conditions where there was corticosteroid release as in conditions of stress, pain, anesthesia, trauma, surgical manipulation etc. Neutrophilia was observed more in group II (control group) for prolonged duration after fracture stabilization, which was also observed by Alam et al. (2005) in case of open fracture repair. On the contrary, neutrophilia was subsided by 14th post-operative day due to use of local antibiotics to defend infection causing organisms.
Based on the values obtained in fracture stabilization of long bone diaphyseal compound fractures in group 1 (with local bone cement-antibiotic incorporation at fracture site) and group II (control group with no local antibiotic therapy) it was concluded that the technique of adopting the use of local antibiotic biodegradable bone cement impregnated LCP and beads in stabilization of compound diaphyseal long bone fractures provided remarkable improvement in the limb function and maintained good implant stability throughout the period of study with haematological and biochemical parameters returning to normal early in the post-operative period compared to control group. Here the infection was controlled by adding gentamicin and the bone healing was facilitated by the use of biodegradable bone cements like hydroxyapatite and Poly DL, lactide. The present study showed that unnecessary long term systemic antibiotic usage for open and infected fractures in dogs may be avoided by using antibiotic biodegradable impregnated implants in future.