Evaluation of Systemic C-reactive Protein as a Systemic Inflammatory Marker in the Blood for Patients Undergoing Minor Oral Surgical Procedures

ABSTRACT Background: C-reactive protein (CRP) is an acute phase protein that its plasma levels increase after trauma or surgery so it is used as an indicator for the level of inflammation after surgery. The objective of this study is to investigate preand post-operative levels of CRP in three types of oral surgical interventions (Apicoectomy, Impaction, and Impacted teeth exposure). Materials and Methods: A total number of (48) healthy individuals aged (20-60) years who needed oral surgical intervention for either (removal of impacted third molars, exposure of an impacted canine, or Apicoectomy). A 4ml venous blood was obtained from each patient at two occasions (pre-operatively at the day of operation and postoperatively after 48 hours), then centrifuged for 15 minutes at (1000x g) and finally the sera were separated and stored at (-20o C) to be used for later analysis by Enzyme Linked Immuno Sorbent Assay (ELISA). Results: There was statistically high significant increase in the level of CRP after oral surgical interventions in all types of operations (the CRP mean value pre-operatively was 2.407925 mg/L, post-operatively after 48hr was 8.486725 mg/L, and the P-value was 0.000). Also there was a high significant difference between apicectomy, impaction, and exposure operations in the level of CRP at day two postoperatively (P-value was 0.000). Conclusion: An inflammatory process develops after oral surgical interventions which necessitate the use of antiinflammatory agents after these procedures and that the severity of inflammation measured by means of CRP levels is correlated to the degree of swelling, length, and type of the surgical procedure.


INTRODUCTION
The common postoperative consequences of the oral surgical interventions are pain and swelling (1) .The acute inflammation that evolves as a result of the surgical handling of the soft and hard tissues is the primary cause of these symptoms and signs (2) .Anti-inflammatory agents are usually prescribed to reduce the postoperative inflammatory sequelaeare produced by the inflammatory process following these procedures (1) .A lot of these anti-inflammatory drugs specially steroidal and non-steroidal antiinflammatory agents have many side effects and are very toxic to body organs particularly to the liver and kidney.According to American Medical Association Media in each year about 20000 patients die from these drugs and another 100000 patients will end up in the hospital due to liver damage, kidney damage, or intestinal bleeding (3) .These complications can be randomly predicted by many factors related with oral surgical procedures such as pre-operative inflammation, pre-operative infection, type of operation, time of operation, technique of operation (4,5) .
(1) Master student.Department of Oral and Maxillofacial Surgery.College of Dentistry, University of Baghdad.
(2) Assistant professor.Department of Oral and Maxillofacial Surgery.College of Dentistry, University of Baghdad.
But these factors alone are not definitive or reliable indicators for the postoperative complications.So the knowledge of reliable indicators of post-operative complications will assess the patient and the operator to manage these complications (6) .The C-reactive protein(CRP) is an acute phase protein of inflammatory status that increases after surgery or any traumatic situation and in a healthy individual the CRP value is less than 10 mg/L (7,8) .The CRP is produced by the liver in reaction to a variety of stimuli that include damaging of tissues.Small amounts of CRP is found in the blood of healthy persons (9,10) .It has been found that the severity of postoperative outcomes are associated with the level of CRP which is a sensitive biomarker of systemic inflammatory status (11) .Estimation of inflammatory indicators has been regarded as a way to improve the anticipation of the risk of these outcomes.The aim of this study is to evaluate the CRP levels pre-and post-operatively as a biomarker for the level of inflammation following minor oral surgical interventions so that to make a prediction for the need of use of anti-inflammatory agents with these interventions.Exclusion Criteria 1-Patients with an active infection (local or systemic).2-Medically and immunologically compromised patients such as diabetic patients, endocrin disturbance, anemia, leukemia tumors or metastatic diseases.3-Patients with social habits such as alcohol consumption or smoking.4-Patients on steroid therapy or oral contraceptives.5-Pregnant and breast feeding women.

Serum Sample Collection
A 4ml venous blood was patientat two occasions (pre day of operation and post hours), then centrifuged for 15 minutes at (1000x g) and finally the sera were separated and stored at (-20º  Subjects without any systemic infection or acute illness Steroids, anti-inflammatory agents should not be taken by the patients in the last week before

Exclusion Criteria
Patients with an active infection (local or systemic).
Medically and immunologically compromised patients such as diabetic patients, endocrin disturbance, anemia, leukemia tumors or metastatic diseases.
Patients with social habits such as alcohol consumption or smoking.
Patients on steroid therapy or oral contraceptives.
Pregnant and breast feeding women.Patients with social habits such as alcohol consumption or smoking.

Serum Sample Collection
Patients on steroid therapy or oral Pregnant and breast feeding women.

Serum Sample Collection
A 4ml venous blood was collected at two occasions (pre-operatively at the day of operation and post-operatively after 48 , then centrifuged for 15 minutes at (1000x g) and finally the sera were separated and stored ) to be used for later analysis Enzyme Linked Immuno Sorbent Assay (ELISA) quantitative determination of sensitive CRP in human serum.
were done under supervision of the specialized at the Teaching Laboratory unit of Baghdad    In case of a usedthrough labial approach.Then the silk suture The l ength of the surgical procedure measured from the starting of the incision to the finishing of the flap suturing.

Laboratory procedure
Microtiterstrips coated with anti antibody are incubated with diluted standard sera and patient samples.During this incubation step CRP is bou mixing containers of the After removal of the unbound serum proteins by a washing procedure, the antigen complex in each well is detected with specific peroxidase conjugated antibodies. of the unbound conjugate, the strips are incubated with a chromogen solution containing tetramethy blue colo of immunocomplex bound to the wells of the strips.The enzyma addition of 2 N H2SO4 and the absorbance values at 450 nm are determined.ength of the surgical procedure measured from the starting of the incision to the finishing of the flap suturing.

Laboratory procedure
Microtiterstrips coated with anti antibody are incubated with diluted standard sera and patient samples.During this incubation step CRP is bound specifically to the wells mixing containers of the After removal of the unbound serum proteins by a washing procedure, the antigen complex in each well is detected with specific roxidase conjugated antibodies. of the unbound conjugate, the strips are incubated with a chromogen solution containing tetramethylbenzidin and hydrogen peroxide, olor develops in proportion to the amount of immunocomplex bound to the wells of the strips.The enzymatic reaction is stopped by the addition of 2 N H2SO4 and the absorbance values at 450 nm are determined.

9: Microplate ELIZA h College Dentistry
Oral and Maxillofacial Surgery and Periodontics flap was sutured by using 3/0 black simple interrupted technique.ength of the surgical procedure measured from the starting of the incision to the finishing of the flap suturing.

Laboratory procedure
Microtiterstrips coated with anti antibody are incubated with diluted standard sera and patient samples.During this incubation step nd specifically to the wells mixing containers of the high sensitive After removal of the unbound serum proteins by a washing procedure, the antigen complex in each well is detected with specific roxidase conjugated antibodies. of the unbound conjugate, the strips are incubated with a chromogen solution containing lbenzidin and hydrogen peroxide, r develops in proportion to the amount of immunocomplex bound to the wells of the tic reaction is stopped by the addition of 2 N H2SO4 and the absorbance values at 450 nm are determined.

Figure
The level than the level of CRP at the day of surgery (pre operatively) in all types of operations which indicated levels and it is highly significant (P and there was a significant levels in each type of operation which were (P value=0.003,P for apio respectively.
The level of CRP at the 2nd day of surgery in each type of operation revealed that the value and the standard deviationwere as follow for Apic Impaction (8.98 Exposure (1.82 (28.55, 22.65, 5.89) for Apic and Exposure respectivel significant difference between impaction, and of CRP at day two post value=0.000).  he level of CRP at the 2nd day was higher than the level of CRP at the day of surgery (pre operatively) in all types of operations which d that there is a difference in the CRP levels and it is highly significant (P and there was a significant levels in each type of operation which were (P value=0.003,P-value=0.000,and P ocectomy, i respectively. he level of CRP at the 2nd day of surgery in each type of operation revealed that the value and the standard deviationwere as follow for Apicoectomy (13.02 Impaction (8.98 mg/L and Exposure (1.82 mg/L and (28.55, 22.65, 5.89) for Apic and Exposure respectivel significant difference between mpaction, and exposure operations in the level of CRP at day two post value=0.000).

and post-operative CRP levels
of CRP at the 2nd day was higher than the level of CRP at the day of surgery (pre operatively) in all types of operations which that there is a difference in the CRP levels and it is highly significant (P and there was a significant difference in CRP levels in each type of operation which were (P value=0.000,and P impaction, and he level of CRP at the 2nd day of surgery in each type of operation revealed that the value and the standard deviationwere as follow ectomy (13.02 mg/L and mg/L and± 4.83); and for mg/L and± 1.59) with mean (28.55, 22.65, 5.89) for Apicoectomy, Impac and Exposure respectively,and there was a high significant difference between xposure operations in the level of CRP at day two post-operatively (P evel of CRP at 2 surgery of CRP at the 2nd day was higher than the level of CRP at the day of surgery (pre operatively) in all types of operations which that there is a difference in the CRP levels and it is highly significant (P-value=0.000)difference in CRP levels in each type of operation which were (P value=0.000,and P-value=0.008)mpaction, and exposure he level of CRP at the 2nd day of surgery in each type of operation revealed that the mean value and the standard deviationwere as follow mg/L and± 6.39); for 4.83); and for ) with mean rank ectomy, Impaction, there was a high significant difference between apicectomy, xposure operations in the level operatively (P evel of CRP at 2 nd day of There was a between the time of surgery (length of the surgical procedure) and the CRP day of operation (correlation coeffi and P value=0.000).
The mean value and the standard deviation the time of surgery Apicoectomy (20.90 Impaction (20.70 Exposure (19.55 ank (30.64, 19.03, 11.39) for Apic Impaction, and Exposure respectively was a significant difference impaction, and exposure operations in the operation (P-value=0.001).

DISCUSSION
The statistical analysis revealed that the levels of CRP post-operatively were higher than pre operatively in all types of operations and there was a high significant difference which indicate surgical intervention in all types of operations.
his is in agreement with al (12) ,Desai (13) and researchers maintained intervention, the level patients.
These findings can be explained by the fact thatthe surgical trauma create a unique metabolic reaction involving a raise in the circulating stress hormones and a raise in the production and secretion of different acute the CRP which blood and its concentration increase in response to inflammatory condition because of it is an acute phase protein.
showed that there is a high significant difference amongApicoectomy, Impaction, an operations in the level of CRP at day two postoperatively and the highest value was found

DISCUSSION
The statistical analysis revealed that the levels operatively were higher than pre operatively in all types of operations and there was a high significant difference which indicated that CRP level increase after surgical intervention in all types of operations.
his is in agreement with and HaoShen maintained intervention, the level of CRP raised in all These findings can be explained by the fact thatthe surgical trauma create a unique metabolic involving a raise in the circulating stress hormones and a raise in the production and secretion of different acute which is located in the plasma of the blood and its concentration increase in response to inflammatory condition because of it is an acute phase protein.The that there is a high significant difference ectomy, Impaction, an operations in the level of CRP at day two postoperatively and the highest value was found The statistical analysis revealed that the levels operatively were higher than pre operatively in all types of operations and there was a high significant difference among CRP level increase after surgical intervention in all types of operations.
his is in agreement withCha HaoShen et al (14) all thos that after surgical of CRP raised in all These findings can be explained by the fact thatthe surgical trauma create a unique metabolic involving a raise in the circulating stress hormones and a raise in the production and secretion of different acute-phase proteins is located in the plasma of the blood and its concentration increase in response to inflammatory condition because of it is an The statistical analysis that there is a high significant difference ectomy, Impaction, and Exposure operations in the level of CRP at day two postoperatively and the highest value was found The statistical analysis revealed that the levels operatively were higher than preoperatively in all types of operations and there among them CRP level increase after surgical intervention in all types of operations.
Chander et all those that after surgical of CRP raised in all These findings can be explained by the fact thatthe surgical trauma create a unique metabolic involving a raise in the circulating stress hormones and a raise in the production and phase proteins like is located in the plasma of the blood and its concentration increase in response to inflammatory condition because of it is an statistical analysis that there is a high significant difference d Exposure operations in the level of CRP at day two postoperatively and the highest value was found Oral and Maxillofacial Surgery and Periodontics 115 in Apicoectomy and the lowest value was found in Exposure.This may be attributed to the more degree of tissue damage and trauma as a result of the more surgical handling of the soft and hard tissues in Apicoectomy procedures ascompared to Exposure procedures or may be due to the longer operation time in Apicoectomycompared to Exposure operation.The statistical analysis showed that there is high significant relationship between the time of operation and the level of CRP at the 2 nd day of operation with all types of operations,and there was highly significant difference amongApicoectomy, Impaction, and Exposure operations in the time consumed in these operations.
These findings are in agreement withMaria et al (15) andSanchis et al (16) who maintained that the postoperative tissue reactions following oral surgery have been commonly related with the length of the surgical intervention.These findings can be explained by the fact that the more operation time means more tissue damage which lead to more elevation in CRP levels.Ohzato et al (17) showed that the magnitude of CRP reaction change instantly with the sharpness of tissue destruction, kind of inflammatory stimulation and type of tissue involved in the condition.Furthermore the longest time was found in Apicoectomy and the shortest time was found in Exposure.
In conclusion this study showed that there was significant increase in the level of high sensitive CRP in the blood of patients after all types of surgical intervention when compared with pre-operative levels.And there were significant differences in the mean levels of high sensitive CRP among different surgical procedures (Apicoectomy, Impaction, and Exposure).This research gives an idea that an inflammatory process develops after oral surgical interventions so we recommend the use of antiinflammatory agents after these surgical procedures to reduce the inflammatory process.The level of high sensitive CRP in the patient's serum appear to be positively correlated with the time of the surgical procedure which increase in the prolonged procedures.

Figure impacted Figure
Figure impacted

Figure 9 h
Figure 9

Figure
Figure

Figure 12 :
Figure 12: Level of CRP at 2 surgery significant correlation between the time of surgery (length of the surgical procedure) and the CRP day of operation (correlation coeffi value=0.000). he mean value and the standard deviation the time of surgery w ectomy (20.90min.and Impaction (20.70 min and Exposure (19.55 min and ank (30.64, 19.03, 11.39) for Apic Impaction, and Exposure respectively was a significant difference and exposure operations in the value=0.001).time of surgery in each operation type between the time of surgery (length of the surgical procedure) and the CRP level at the 2nd day of operation (correlation coefficient=0.560 he mean value and the standard deviation were as follow for min.and± 9.62); for min and± 12.15); and for min and± 14.08) with mean ank (30.64, 19.03, 11.39) for Apic Impaction, and Exposure respectively was a significant difference amongapicectomy, and exposure operations in the time of surgery in each operation between the time of surgery (length of the at the 2nd cient=0.560 he mean value and the standard deviation of ere as follow for 9.62); for 12.15); and for ) with mean ank (30.64, 19.03, 11.39) for Apicoectomy, and there picectomy, and exposure operations in the time of time of surgery in each operation

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