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ORIGINAL RESEARCH  
Year : 2010  |  Volume : 21  |  Issue : 4  |  Page : 486-490
Estimation of salivary amylase and total proteins in leukemia patients and its correlation with clinical feature and radiographic finding


Department of Oral Medicine and Radiology, Bapuji Dental College and Hospital, Davangere, India

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Date of Submission24-Jul-2009
Date of Decision06-Jan-2010
Date of Acceptance22-Jul-2010
Date of Web Publication24-Dec-2010
 

   Abstract 

Background: Leukemia is a fatal disease. The oral manifestations of the leukemias occur early in the course of the disease and these oral features can at times act as a diagnostic indicator. Saliva has been used as a diagnostic aid in a number of systemic diseases.
Materials and Methods: In our study, samples of unstimulated saliva of 30 leukemia patients who were not on chemotherapy were collected and analyzed for salivary amylase and total protein. The oral manifestations and radiographic changes (OPG) were recorded. The correlation between the oral manifestations and the salivary components (salivary amylase and total protein) was assessed for prognostic significance.
Results: In the present study when the mean values of salivary amylase (1280±754 U/ml) and total protein (647.2±320.7 mg%) were compared with that in control subjects. There was a statistically significant difference for amylase levels (P<.05). On intraoral examination the study subjects showed pallor, gingivitis, gingival enlargement, petechiae, and ecchymosis. On the OPG, the radiographic features included generalized rarefaction of bone (20%), thinning of lamina dura (3.4%), generalized alveolar crest bone resorption (30%), thinning of walls of alveolar crypts (6.7%), besides others, e.g., periapical abscess (10%).
Conclusions: The saliva of leukemic patients demonstrated obvious changes in composition. A rise in salivary amylase and total protein levels was evident, with the increase in amylase levels being statistically significant.

Keywords: Leukemia, salivary amylase, salivary total protein

How to cite this article:
Ashok L, Sujatha G P, Hema G. Estimation of salivary amylase and total proteins in leukemia patients and its correlation with clinical feature and radiographic finding. Indian J Dent Res 2010;21:486-90

How to cite this URL:
Ashok L, Sujatha G P, Hema G. Estimation of salivary amylase and total proteins in leukemia patients and its correlation with clinical feature and radiographic finding. Indian J Dent Res [serial online] 2010 [cited 2019 Oct 22];21:486-90. Available from: http://www.ijdr.in/text.asp?2010/21/4/486/74212
Many diseases that involve the whole body produce oral changes at some stage in their course. These oral manifestations are often mild and overshadowed by the major manifestations of the systemic disease. [1] Leukemia is a uniformly fatal disease of unknown etiology that is characterized by excessive and abnormal proliferation of primitive white blood cells and their precursors, with infiltration of the various tissues of the body, especially bone marrow, spleen, and lymph nodes. [2]

It has been recognized that some patients with acute leukemia will first consult a dentist because of the oral manifestations of their underlying disease. Early reports in the literature urge dentists to be aware of the possible oral manifestations of leukemia so to avoid the potentially catastrophic post-surgical complications of oral hemorrhage, tissue necrosis, and infection. [3]

The most commonly used laboratory diagnostic procedures involve the analysis of the cellular and chemical constituents of blood. Other biologic fluids can also be utilized for the diagnosis of disease, and saliva offers some distinctive advantages over serum because it can be collected noninvasively by individuals with modest training. Diagnosis of disease via the analysis of saliva is especially valuable in children and older adults and may provide a cost-effective approach when screening large populations. [4]

Salivary amylase and total protein content in the saliva of leukemia patients are found to be elevated and therefore can be of diagnostic value. [5]

Diagnostic imaging is interpreted along with clinical findings to yield a diagnosis. In addition, radiographs may be of value to identify clinically undetectable pathosis. The use of panoramic radiography to screen patients with cancer has received some attention in the literature. The information yield from panoramic radiographs and their effectiveness as a diagnostic tool needs to be examined more carefully, particularly in patients with leukemia in whom the risk of infection is great. [6]

The present study was undertaken to find out whether any correlation exists between the severity of oral manifestations and the bony changes seen on orthopantomograph (OPG) with the salivary amylase and total protein in leukemia patients.

Clinical manifestations

Since all leukemias disrupt hematopoiesis, it is not surprising that all forms of leukemia have some features in common. The clinical features of acute and chronic leukemias reflect the different rates of leukemic cell accumulation and the degree of bone marrow function inhibition. [7],[8],[9],[10]

The clinical manifestations include, fever, pallor, weakness and loss of weight, anemia, bleeding manifestations, infections, lymphadenopathy, hepatosplenomegaly, pain, and chloromas. The oral manifestations include, pallor, gingivitis, gingival enlargement, petechiae, and ecchymosis. Cooper et al., [11] reported a case of 35-year-old male patient of acute myeloid leukemia (AML) and observed that gingival hyperplasia was most commonly seen with the AML subtypes of acute monocytic leukemia (M5) (66.7%), acute myelomonocytic leukemia (M4) (18.5%), and acute myelocytic leukemia (M 1 , M 2 ) (3.7%). Wu et al., [12] reported a case of 53-year-old male patient with AML who presented with gingival enlargement, fatigue, and recent weight loss as the initial manifestations of acute myelomonocytic leukemia. They stressed that the most frequently observed oral findings in leukemia are mucosal bleeding, ulceration, petechiae, and gingival hyperplasia.

The radiographic manifestations in the jaws in leukemia include generalized rarefaction of bones, thinning of the lamina dura, alveolar crest bone resorption, and periapical lesions.

Felix and Lukens [13] stressed the importance of the identification of acute gingival hypertrophy for the early recognition of acute nonlymphocytic leukemia. They stated that dental and medical personnel should be alert to these findings, as the oral signs and symptoms may be characteristic of serious systemic diseases. Morgan [14] presented a case of 77-year-old female patient who complained of a nodular swelling of the gingiva in relation to the mandibular premolar, with a vital pulp and a periapical radiolucent lesion; she was diagnosed with chronic lymphocytic leukemia (CLL). They stated that this infiltrative lesion in the jaw was an indication of a highly aggressive CLL that soon resulted in the appearance of thrombocytopenia.

Investigations and diagnosis of leukemias

The diagnosis of leukemia may be suspected on clinical grounds but is confirmed only by finding the characteristic morphological, cytochemical, and immunohistochemical features on examination of blood and bone marrow aspirates. In problematic cases resort to cytogenetics and molecular biology may be necessary. As a diagnostic fluid, saliva offers distinctive advantages over serum because it can be collected noninvasively by individuals with modest training. Furthermore, saliva may provide a cost-effective approach for the screening of large populations. [4]

Rahematulla et al., [5] analyzed the salivary constituents in AML and acute lymphoblastic leukemia (ALL) patients and reported that the mean values for amylase activity and total protein concentrations were significantly higher in these subjects. They assessed several salivary components in stimulated whole saliva from patients with acute leukemia who were undergoing chemotherapy. Saliva samples were collected at the time of diagnosis and longitudinally during the treatment period. Data analysis showed that patients with leukemia had significantly higher peroxidase and amylase activity and elevated concentration of salivary total protein at the time of diagnosis.


   Materials and Methods Top


The present study group comprised 30 patients visiting the General Medicine Department. The control group consisted of 25 age- and sex-matched healthy subjects [Table 1], [Table 2] and [Table 3] for salivary analysis and 10 controls for radiographic examination [Table 4]. Salivary total protein was estimated by using ERBA ChemPro, semiautomated analyzer (Transasia Bio-Medicals Ltd., Mumbai) and two reagents (Autospan reagents, Span Diagnostics Ltd., Surat, India). Reagent 1 included Biuret reagent 1×100 ml, copper sulphate, sodium hydroxide, sodium potassium tartarate, and surfactant. Reagent 2 included protein standard 6.5 gm/dl, 1×3 ml BSA, buffer, and stabilizer.
Table 1: Composition of the control group for salivary analysis

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Table 2: Disease distribution according to age

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Table 3: Disease distribution according to sex

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Table 4: Composition of the control group for radiographic examination

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Salivary amylase was estimated by using an autoanalyzer (CoBAS Integra; -400 fully-automated biochemistry analyzer, Germany). Materials used are reagents (Roche Cobas Integra; Reagent, a-amylase - CNP-G3 (2-chloro-4-nitro a maltotrioside).

Methodology

Patients with a diagnosis of leukemia and who were not on chemotherapy were eligible for inclusion in the study. Saliva sample of all the study subjects were collected in the morning between 8.30 AM and 9.00 AM. Unstimulated saliva was collected from the floor of mouth after 5 minutes using a sterile disposable cartridge and transferred to a sterile collecting tube. Two milliliters of saliva was kept immediately in the refrigerator for further analysis.

Control group

Twenty-five age- and sex-matched healthy subjects were selected as controls for salivary analysis and 10 subjects in the age-group of 5-65 years (mean age 35 years) for radiographic study. The control subjects did not suffer from any systemic diseases that could affect the properties of their saliva.

Estimation of salivary total protein was done by the modified biuret method. [15] Estimation of salivary amylase was done using the CNP-G 3 method. [16],[17] The procedure was as follows: The samples and the reagent were brought to room temperature prior to use. About 1 ml of reagent1 and 1.5 μl of sample was dispensed into a test tube, mixed, and read immediately using a fully-automated analyzer. The values displayed on monitor were recorded and entered in a proforma.

Statistical analysis

Descriptive data are presented as mean±SD. One-way ANOVA was used for multiple group comparisons followed by the Mann-Whitney test for assessing the difference between groups. P≤.05 was considered as indicating statistical significance.


   Results Top


The present study aimed to analyze selected parameters in the saliva of 30 leukemia patients who were not on chemotherapy and to find out if there was any correlation between these parameters and the oral manifestations and radiological changes (in orthopantomographs) of these patients. Out of 30 subjects, 13 (43.4%) had AML, 7 (23.4%) had ALL, 6 (20%) had CML, and 4 (13.33%) had CLL. The age of the study subjects varied from 3 to 80 years, with a mean age of 41.5 years. Out of 30 study subjects, 20 were males and 10 were females. The AML group consisted of 7 males and 6 females, the ALL group consisted of 5 males and 2 females, the CML group consisted of 4 males and2 females, and the CLL group of 4 males.

Salivary total protein


Among the 25 control subjects, the minimum level of salivary total protein was 180 mg% and the maximum was 1320 mg% (mean: 594 mg%) [Table 1]. The normal range of total protein in saliva is reported to be 140-640 mg%. [18] Out of 30 experimental subjects, 16 (53.4%) showed increased salivary total protein, while 14 (46.7%) had salivary total protein in the normal range. In the study group, the minimum level of salivary total protein observed was 24.7 mg% and the maximum was 1520 mg% (mean: 647.2 mg%) [Table 1]. In acute leukemias the mean value of total protein was 621.09 mg% and in chronic leukemias the mean value was 699.7 mg% [Table 2].

Salivary amylase

After considering the dilution factor (1:10), salivary amylase concentration values were reevaluated for statistical analytical purpose. Among the 25 control subjects, the minimum level of salivary amylase level was 113 U/ml and the maximum was 1125 U/ml, with the mean being 737 U/ml [Table 1]. All the subjects showed normal salivary amylase levels; the normal values range from 27±3.8 to 1440±160 U/ml. (The wide variation in the concentration of α-amylase could be due to differences in the assay techniques, the reagents used, glandular contribution, and carbohydrate consumption).[19]

Out of the 30 experimental subjects, 5 (16.7%) (1 ALL, 1 CML, and 3 CLL patients) showed increased salivary amylase levels, while 25 (83.4%) had salivary amylase in normal range. The minimum level of salivary amylase observed in study group was 512 U/ml and the maximum was 3940 U/ml [Table 1]. The mean value was 1280 U/ml. In acute leukemias, the mean value of salivary amylase was 1061 U/ml, and in chronic leukemias the mean was 1718 U/ml [Table 2].

Oral manifestations

None of the 25 control subjects showed any mucosal lesions. Out of the 30 experimental subjects, 25 (83.4%) had pallor (12 AML, 7 ALL, 5 CML, and 1 CLL patients); 10 (33.4%) had gingivitis (4 AML, 2 CML, and 4 CLL patients); 16 (53.4%) had gingival enlargement (4 AML and 1 ALL patients); 16 (53.33%) had enlarged, palpable, and tender submandibular lymphnodes (5 AML, 4 ALL, 3 CML, and 4 CLL patients); 5 (16.7%) had petechiae (3 AML and 2 ALL patients); and 1 patient (3.4%) had ecchymosis.

Radiographic features

Among the 20 patients with acute leukemias, 3 (1.5%) had generalized rarefaction of bone, 6 (30%) had generalized alveolar crest bone resorption, 2 (10%) had thinning of the walls of the alveolar crypts, and 2 (10%) showed periapical lesions. Among the 10 patients with chronic leukemias, 3 (30%) had generalized rarefaction of bone, 1 (10%) had generalized thinning of the lamina dura, 3 (30%) had generalized alveolar crestal bone resorption, and 1 (10%) patient showed a periapical lesion (periapical radiolucency with widening of the PDL and loss of the lamina dura were considered as 'abscess/periapical lesion' for the purpose of this study).

Comparison of healthy controls and leukemia subjects

Salivary total protein: The mean salivary total protein was 594.0±353.3 for the controls (the normal range is 140-640 mg%), while the study subjects had a mean value of 647.2±320.7. This difference was nonsignificant (P=.43) [Table 5].
Table 5: Salivary total protein and salivary amylase levels

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The mean salivary total protein was 594.0±353.3 among the controls, whereas the mean values in acute and chronic leukemia patients were 621.0±274.7 and 699.7±409.2, respectively. The difference from the control values was not statistically significant (P=.49 and P=.55, respectively) [Table 6].
Table 6: Comparison of salivary total protein and salivary amylase

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Salivary amylase: The mean salivary amylase in the control group was 737±301, whereas it was 1280±754 in the study subjects; this difference was statistically significant (P<.05). The normal range of salivary amylase ranges from 27±3 to 144±160 U/ml [Table 5].

The mean value of salivary amylase in the control group was 737±301 U/ml as compared with the mean values of 1061±353 U/ml and 1718±1118 U/ml in the acute and chronic leukemia groups, respectively. The difference from the control group was statistically significant (P<.01) [Table 6].


   Discussion Top


Oral manifestations in all types of leukemias occur early in the course of the disease and many a time act as a diagnostic indicator in leukemias.

Leukemia is a disease of the hematopoietic tissues and affects the bone marrow. Thus diagnostic imaging can provide additional information that can aid in arriving at a diagnosis. The use of panoramic radiography to screen patients with cancer has received some attention in literature and its effectiveness as a diagnostic tool needs to be examined more carefully, particularly in patients with leukemia in whom the risk of infection is great. [9]

It is becoming increasingly apparent to investigators and clinicians in a variety of disciplines that saliva has many diagnostic uses and can be especially valuable in large-scale screening and epidemiologic studies. Saliva is being used as a diagnostic aid in an increasing number of clinical situations in systemic diseases that can affect salivary gland function and the composition of saliva. [20]

This study was designed to gain further insights into the composition of saliva in leukemia patients and the potential for using saliva as a screening tool in the diagnosis of leukemias.

Oral manifestations

In the present study, 25 patients (83.4%) had oral manifestations in the form of pallor (82.4%), gingivitis (33.4%), gingival enlargement (16.7%), submandibular lymphnode enlargement (53.4%), and petechiae and ecchymosis (16.7%). This corresponds well with the findings of Duffy and Driscoll, [21] where 80% of patients showed oral manifestations, with gingivitis in 55%, gingival hypertrophy in 45%, bleeding in 42%, ulcers in 39%, and petechiae in 7%. Our findings are also consistent with the findings of Barrette [22] who observed oral signs in 77% of the patients.

Radiographic features

In 30 study subjects, 6 (20%) had generalized rarefaction of bone, 1 (3.4%) had thinning of the lamina dura, 9 (30%) had generalized alveolar crest bone resorption, 2 (6.7%) had thinning of walls of alveolar crypts, and 3 (10%) showed periapical lesions.

Salivary total protein and amylase

In the present study, the mean values for salivary amylase in the study subjects was 1280±754, which was significantly different from the value seen in control subjects (P<.05). The mean salivary total protein of study subjects was 647.2±320.7 compared with the mean of 594.0±353 in the control group; this difference was statistically nonsignificant (P=.43).

A rise in salivary total protein in whole saliva is also seen in other patients, including those with newly diagnosed untreated oral and phayrngeal cancer, diabetes, cystic fibrosis, and stomatitis secondary to chemotherapy, etc. The cause of this increase in salivary protein and amylase concentrations in patients with certain diseases is not known.


   Conclusion Top


In conclusion, in the present study the saliva of leukemic patients demonstrated obvious changes in composition. A rise in salivary amylase and total protein levels was evident, with the increase in amylase levels being statistically significant. However, it must be kept in mind that whole saliva is a complex mixture of glandular secretions, oral microorganisms, epithelial cells, erythrocytes, polymorphonuclear leukocytes, and food debris, and these changes are not specific. Increase in salivary protein has also been reported in diseases like untreated oral and pharyngeal cancer, diabetes, and stomatitis secondary to chemotherapy.

This study revealed oral manifestations in most of the patients. The OPG of some patients revealed generalized rarefaction, alveolar crest bone resorption, loss of crypt walls, and periapical lesions. Thus, when interpreted in the light of the history and examination findings, panoramic radiographs of patients with leukemia, can help in arriving at a diagnosis.

Additional and more detailed biochemical analysis on a larger group of patients is needed. In this study it was difficult to have the same number of age-matched patients in each group due to the nature of disease. We included all patients with newly diagnosed leukemia and the number of patients in each group could not be controlled. Findings from further studies on larger groups may justify the use salvia as a prognostic indicator in such patients.

 
   References Top

1.Henderson ES, Lister TA. Leukemia. 5 th ed. Philadelphia: W.B. Saunders company; 1990.   Back to cited text no. 1
    
2.Anil S, Lakshman P, Samarayanake IP, Nair RG, Beena VT. Gingival enlargement as a diagnostic indicator in leukemia. Case report. Aust Dent J 1996;41:235-7.   Back to cited text no. 2
    
3.Barrette AP. Oral Changes as initial diagnostic indicators in acute leukemia. Oral Med 1986;41:234-8.   Back to cited text no. 3
    
4.Kaufman E, Lamster IB. The diagnostic applications of saliva - A review. Crit Rev Oral Biol Med 2002;13:197-212.   Back to cited text no. 4
[PUBMED]  [FULLTEXT]  
5.Rahematulla BM, Techanitiswad I, Rahematulal F, Tonya O, McMillon, Bradley EL. Analysis of salivary components in leukemia patients receiving chemotherapy. Oral Surg Oral Med Oral Pathol 1992;73:35-46.   Back to cited text no. 5
    
6.Epstein JB, Guiseppe R. The value of panoramic radiographic examination in patient with leukemia before medical management. Oral Surg Oral Med Oral Pathol 1992;74:736-41.   Back to cited text no. 6
    
7.Isselbacher KJ, Braunwald E, Wilson JD. Harrison's Principles of Internal Medicine. 13 th ed. New York: Mcgraw Hill Inc;1994.   Back to cited text no. 7
    
8.Edwards CR, Bouchler IA, Haslett C. Davidson's Principles and Practice of Medicine. 17 th ed. Edinburgh: Churchil Livingstone; 1998.   Back to cited text no. 8
    
9.Steron JH, Eisenberg JM, Hertton JJ, Kippel JH. Internal Medicine. New York: Mosby Co.;1998.   Back to cited text no. 9
    
10.Lee GR, Foester J, Lukens J. Wintrobe's Clinical Hematology., 10 th ed, Baltimore: Williams and Wilkins;1998.   Back to cited text no. 10
    
11.Cooper CL, Loewen R, Shore T. Gingival hyperplasia complicating acute myelomonocytic leukemia. J Am Dent Assoc 2000;66:78-9.   Back to cited text no. 11
    
12.Wu J, Fantasia JE, Kaplan R. Oral manifestations of acutemyelomonocytic leukemia: A case report and review of the classification of leukemias. J Periodontol 2002;73:664-8.   Back to cited text no. 12
[PUBMED]  [FULLTEXT]  
13.Felix DE, Lukens J. Oral symptom as a chief sign of acute monoblastic leukemia: Report of case. J Am Dent Assoc 1986;113:899-900.   Back to cited text no. 13
[PUBMED]    
14.Morgan LA. Infiltrate of chronic lymphocytic leukemia appearing as periapical radiolucent lesion. J Endodontics 1995;21:475-8.   Back to cited text no. 14
    
15.Vatzidis H. An improved biuret reagent. Clin Chem 1977;23:908.   Back to cited text no. 15
    
16.Winn Dean ES, David H, Sigler G, Chavez R. Development of a direct assay for a-amylase. Clin Chem 1988;34:2005-8.   Back to cited text no. 16
    
17.Lorentz K. An ideal substrate for the measurement of pancreatic amylase? Clin Chem Lab Med 2002;40:781-5.   Back to cited text no. 17
[PUBMED]  [FULLTEXT]  
18.Jenkins GN. Physiology and Biochemistry of the Mouth. 4 th ed. Oxford: Blackwell Scientific Publications; 1978.  Back to cited text no. 18
    
19.Makinen KK. Salivary enzymes. Chapter 3. Human saliva. Clinical Chemistry and Microbiology. Vol. 2. Flordia: Tenovuo J.O. C.R.C. Press; 1989. p. 91-119.   Back to cited text no. 19
    
20.Mandel ID. The diagnostic uses of saliva. J Oral Pathol Med 1990;19:119-25.   Back to cited text no. 20
[PUBMED]    
21.Duffy JH, Riscoll EJ. Oral manifestations of leukemia. Oral Surg 1958;11:484.   Back to cited text no. 21
[PUBMED]    
22.Barette A. Oral changes as initial diagnostic indicators in acute leukemia. J Oral Med 1986;41:234-8.  Back to cited text no. 22
    

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Correspondence Address:
G Hema
Department of Oral Medicine and Radiology, Bapuji Dental College and Hospital, Davangere
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0970-9290.74212

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    Tables

  [Table 1], [Table 2], [Table 3], [Table 4], [Table 5], [Table 6]

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