Value of Neutrophil (Lymphocyte Ratio), Red Blood Cell Distribution Width, Mean Platelet Volume and C - Reactive Protein in the Assessment of Major Depressive Disorder

  

    
Test Result    Variables
    
AUC
    
p valuea
    
95% CI of AUC
  
  

    
Lower Bound
    
Upper Bound
  
  

    
RDW CV (%)
    
0.819
    
0.001s
    
0.746
    
0.892
  
  

    
MPV (fl)
    
0.949
    
0.001s
    
0.917
    
0.982
  
  

    
NLR
    
0.914
    
0.001s
    
0.870
    
0.959
  
  

    
CRP (mg/L)
    
0.892
    
0.001s
    
0.839
    
0.944
  

Table 5: Area under the curve of RDW-CV, MPV, NLR and CRP by MDD Group (n=130).

Inclusion criteria:

• The patients were included as MDD according to the criteria’s of the Diagnostic and Statistical Manual of Mental Disorders (5th edition) and patients were diagnosed by expert psychiatric clinician
• The healthy controls with no previous psychiatric diseases
• Age was ≥18 years old
• Both sexes

Exclusion criteria:

• The patients were excluded with-
• Anemia
• Active infections, a leukocyte value suggesting infection
• Other inflammatory or autoimmune diseases
• Severe systemic diseases such as - epilepsy, diabetes mellitus, hypertension, cardiac diseases, hepatic or renal failure
• Alcohol or other substances addiction
• Severe head trauma or mental retardation
• Pregnancy
• Vitamin or fish oil intake
• Heavy smoking ( >30cigarettes/day)
• Obesity (BMI > 30 kg/m²) and concomitant drug use for any reason

Total sample size: 130

Group I (MDD group): 75 patients with MDD were evaluated and 65 newly diagnosed MDD patients were enrolled in this study as cases.

Group II (control group): 65 age sex matched healthy subjects were enrolled in this study as controls.

Sampling technique: Consecutive sampling technique was applied for enrollment of the patients in this study. As per inclusion criteria the patients were enrolled in this study. The procedures were explained to the participants and informed written consent was taken (Tables 6,7). Prior to the study pre-testing was carried out among patients equivalent to 10% of total study population to test the applicability of the methodology including the research instruments. Some modification in the questions was made out and was finalized.



Table 6: Distribution of the study population according to groups with RDW-CV (n=130).

  

    
RDW CV (%) 
    
Case    (n=65) 
    
Control    (n=65) 
    
p value*
  
  

    
 
    
n
    
%
    
n
    
%
    
 
  
  

    
Raised (>13.45)
    
48
    
73.8
    
18
    
27.7
    
0.001s
  
  

    
Normal
    
17
    
26.2
    
47
    
72.3
  
  

    
Total 
    
65
    
100
    
65
    
100
    
 
  

Table 6: Distribution of the study population according to groups with RDW-CV
(n=130).

Close



Table 7: Distribution of the study population according to groups with MPV (n=130).

  

    
MPV (fl) 
    
Case    (n=65) 
    
Control    (n=65) 
    
p value*
  
  

    
 
    
n
    
%
    
n
    
%
    
 
  
  

    
Raised (>10.25)
    
58
    
89.2
    
12
    
18.5
    
0.001s
  
  

    
Normal
    
7
    
10.8
    
53
    
81.5
  
  

    
Total 
    
65
    
100
    
65
    
100
    
 
  

Table 7: Distribution of the study population according to groups with MPV
(n=130).

Close

Specimen collection: About 4.0 ml venous blood was collected from each patient through an aseptic venipuncture from antecubital vein. 2.0 ml blood was collected in EDTA tube for Complete Blood Count (CBC) which was measured by haematology auto analyzer (SYSMEX-XT 4000i) and a smear was made for Peripheral Blood Film (PBF) examination. The EDTA containing blood sample was tested within 2 hours of collection. Rest 2.0 ml blood was collected in plain test tube for CRP measurement. CRP was measured by Biochemistry auto analyzer (Dimension RxL Max) within 2 hours of collection.



Table 8: Distribution of the study population according to groups with NLR (n=130).

  

    
NLR 
    
Case    (n=65) 
    
Control    (n=65) 
    
p value*
  
  

    
 
    
n
    
%
    
n
    
%
    
 
  
  

    
Raised (>1.98)
    
55
    
84.6
    
17
    
26.2
    
0.001s
  
  

    
Normal
    
10
    
15.4
    
48
    
73.8
  
  

    
Total 
    
65
    
100
    
65
    
100
    
 
  

Table 8: Distribution of the study population according to groups with NLR
(n=130).

Close

Test procedure: Complete blood count (CBC): CBC was estimated by hematology auto- analyzer (SYSMEX-XT 4000i) which again rechecked manually by peripheral blood film. NLR was measured by dividing neutrophil count to lymphocyte count from CBC report and RDW and MPV was simultaneously reported as a part of CBC.

NLR= Total Neutrophil Count/Total Lymphocyte Count

RDW-CV= 1 SD/MCV×100%

MPV (fl) = Plateleterit (%)/PLT (×10⁹/L)

Peripheral Blood Film (PBF): For examination of peripheral blood film, uniform blood smear was made on a glass slide with the help of spreader. Leishman’s staining was done by standard technique. It was examined by binocular light microscope at first 10x objective then 40x objectives.

C - reactive protein: CRP was measured by Biochemistry auto analyzer (Dimension RxL Max) using the C-Reactive Protein Extended Range (RCRP) method. The RCRP method is based on a particle enhanced turbidimetric immunoassay (PETIA) technique.

Safety precaution: Universal precaution was obtained. Gloves, laboratory coat, and safety glasses was worn when handling all human blood products. Disposable plastic, glass, paper and gloves that contact blood were placed in a biohazard bag. Non-disposable material at the end of working day was disinfected. Washing hands thoroughly was done after removal of personal protective devices used in handling specimens and kit reagents.

Data collection: Data were collected by a pre designed proforma. Blood samples were obtained from the patient. Patient’s information was obtained through using patient’s information sheet which involves, questionnaire, clinical finding & laboratory reports. Statistical Package for Social Science (SPSS) version 23 was used for all statistical analysis. The significance of mean difference between case and control groups were assessed by Student’s t test and Chi-square test was used to compare categorical variables. Data were presented as mean ± SD. Receiver Operating Characteristics (ROC) curve graphics were used in the comparison of sensitivity and specificity. P-values less than 0.05 were regarded as significant.

Results

Among the 12 parameters the mean difference between cases and controls, five parameters shows non-significant result (p>0.05) but rest seven parameters show statistically significant (p<0.05). In this study, Hb, ESR, RBC count, WBC count, PLT count, MCV, MCH, MCHC, Neutrophil%, ANC, Lymphocyte% and ALC were linked to main outcome variables but not direct outcome variables. So these parameters were placed in separated.

Discussion

Among the psychological conditions, depression is a common, complex disease. Depression is associated with prominent disability, social burden and reduced quality of life [11]. Immune response and inflammation play a significant role in the etiology of MDD [12]. This case-control study was conducted in the Department of Laboratory Medicine (Clinical Pathology) in collaboration with the Department of Psychiatry, BSMMU, Dhaka. In this study, Level of NLR, RDW, MPV and CRP were measured in newly diagnosed Major Depressive Disorder patients and compared with healthy controls. This study included total 130 patients of which Group I, 65 patients with MDD who were clinically diagnosed and Group II, 65 controls who did not have any symptom or sign of MDD and were apparently healthy. In this study, it was found that mean age of patients with Group I (MDD) and Group II (control) was 33.06 ± 12.23 and 33.05 ± 11.43 years respectively. The difference of mean age was not statistically significant among two study groups (p>0.05). This observation is consistent with the findings of others [6,12,13]. Demir and co-investigators found that the median age of MDD group and control group was 28.4±9.2 and 30.0±9.2 years respectively. The difference in age between the groups were not also statistically significant (p>0.05).The present study showed that male were 22 (33.8%) and female were 43 (66.2%). Predominance female patient was also found in other studies [6,13,14]. Their study found that male and female ratio was approximately 1:2. These findings were nearly consistent with our study. In this study, it was found that mean Hb were 12.70±0.89 g/dl in cases and 13.09±1.04 g/dl in controls (p<0.05). Mean ESR were 21.35±7.56 mm in 1st hour in cases and 14.52±5.54 mm in 1st hour in controls (p<0.05). Mean RBC count were 4.36±0.54 x 1012/L in cases and 4.52±0.43 x 1012/L in controls (p>0.05). Mean WBC count were 8.87±1.25x109/L of blood in cases and 8.34±1.07 x109/L of blood in controls (p<0.05).Mean PLT count were 302.94±41.14 x109/L of blood in cases and 316.57±47.25 x109/L of blood in control (p>0.05). Mean MCV were 85.61±4.64 fl in cases and 85.37±4.27 fl in control (p>0.05). Mean MCH were 28.62±1.36 pg in cases and 29.04±1.19 pg in control group (p>0.05). Mean MCHC were 33.42±1.21 g/dl in cases and 33.79±1.26 g/dl in control group (p>0.05). Mean neutrophil (%) were 65.63±3.63 in cases and 59.15±3.08 in control group (p<0.05). Mean ANC were 5.83±0.93 x 109/L in cases and 4.94±0.70 x 109/L in control group (p<0.05). Mean lymphocyte (%) were 28.26±3.05 in cases and 33.78±3.07 in control group (p<0.05). Mean ALC were 2.50±0.39 x 109/L in cases and 2.82±0.44 x 109/L in control group (p<0.05). Here, RBC count, PLT count, MCV, MCH and MCHC of mean difference between cases and controls show not statistically significant (p>0.05). These findings are similar to several studies [6,9]. But others like Hb, ESR, WBC count, Neutrophil%, ANC, Lymphocyte%, ALC are statistically significant (p<0.05). These results are consistent with other studies [6,9,13]. These parameters are part of CBC and are linked to the main outcome variables. RDW are derived from the formula of [(one Standard deviation of red cell volume ÷ mean cell volume) x 100] and NLR also derived from dividing absolute neutophil count by absolute lymphocyte count [6]. In our study, it was observed that the baseline NLR were higher in MDD patients than healthy controls. In this study mean NLR in Group I was 2.36±0.38, whereas it was 1.77±0.24 in Group II. The differences between the Group I (case) and Group II (Control) of NLR were statistically significant (p<0.001). These results agreed to the findings of other studies [6,13]. Demircan F et al. [6] found that mean NLR was 2.55±0.70 in patients with MDD, whereas it was 1.41±0.8 in control group (p<0.001). Peng YF et al., [13] also found higher mean NLR in patients with MDD group compared to the control group (1.9±0.73 vs. 1.7±0.62) (p<0.05). In this study, mean RDW-CV (%) level was found to be 14.20±1.08 in patients with MDD and mean RDW-CV (%) level was found to be 13.03±0.61 in the control group. RDW levels were significantly higher in patients with MDD compared to the control group (p<0.001). These findings are similar to a study conducted by Demircan F et al., [6]. They also found higher mean RDW in patients with MDD group compared to the control group (14.3±2.6 vs. 13.4±1.8) (p<0.001). Peng YF et al., [13] also found higher mean RDW in patients with MDD group compared to the control group (13.4±0.87 vs. 12.9±0.58) (p<0.001). These findings are also consistent with present study. In the present study, it was found that means MPV level was 10.93±0.54 fl in patients with MDD group and mean MPV level was 9.39±0.91 fl in the control group. There was high MPV level in patients with MDD when compared with controls. The differences between the patients with MDD group and control group of MPV were statistically significant (p <0.001). These results are consistent with other studies [6,14]. Demircan F et al., [6] found higher mean MPV in patients with MDD group compared to the control group (10.4±1.46 vs. 9.33±1.14) (p<0.001). Ataoglu A et al., [14] also found higher mean MPV in patients with MDD group compared to the control group (10.92±0.75 vs. 10.01±0.66) (p<0.001). This study demonstrated that mean CRP level was 5.25±1.84 mg/L in patients with MDD group and mean CRP level was 2.56±1.16 mg/L in the control group. CRP levels were significantly higher in depression group relative to controls (p<0.001). The present study is comparable with several studies that have reported a positive association between CRP and depression [10,15,17]. Sawyer J., [14] showed mean CRP levels were significantly higher in the depressed population than in the healthy control group (4.4±8.4 mg/L vs. 1.6±2.0 mg/L) (p<0.05). Jangpangi D et al., [10] also found that the serum levels of CRP (ng/ml) were significantly higher in depression group as compared to the control group (2132±50.24 vs. 1969±69) (p<0.05). Sensitivity and specificity of NLR, RDW, MPV and CRP were determined by Receiver-Operator Characteristic (ROC) curves (Tables 8-10). The curves also determines about prediction of MDD. In ROC analysis, a cut point of 1.98 was identified for NLR in Group I (case). Area under curve was 0.914, 95% CI of AUC was 0.870 - 0.959 for NLR. An NLR value of more than 1.98 demonstrates a sensitivity of 84.6% and a specificity of 73.8%. This study findings correlate with a similar study conducted by Sunbul E A et al., [7]. In ROC curve, they found a cut of point of 1.57 in MDD patients and concluded that an NLR value ≥ 1.57 revealed a sensitivity of 61.4% and a specificity of 61.2%. In another study conducted by Demircan F et al., [6] also found similar result. The cut off value for NLR was 2.05 for diagnosis of MDD in that study population. ROC analysis showed that an optimal RDW cut off value was 13.45% in patients with MDD. Area under curve was 0.819, 95% CI of AUC was 0.746- 0.892 for RDW. A RDW value ≥ 13.45% shows a sensitivity of 73.8% and a specificity of 72.3% for assessing MDD patients. These results are consistent with other studies [6,11]. Peng Y F et al., [11] showed that an optimal RDW cut off value was 13.05% (area under the curve 0.780, 95 CI%: 0.729-0.831) (p<0.001) in patients with MDD, the sensitivity and specificity of an elevated RDW values were 75.69% and 70.66%, in assessing MDD patients. In ROC analysis, a cut off value of 10.25 fl was found for MPV in Group I (case). Area under curve was 0.949, 95% CI of AUC was 0.917- 0.982 for MPV. A MPV value ≥ 10.25 fl demonstrates a sensitivity of 87.7% and a specificity of 73.8% in assessing MDD patients. Similar result is also found by other studies [6,13]. ROC analysis showed that an optimal CRP cut off value was 3.58 mg/L in patients with MDD. Area under curve was 0.892, 95% CI of AUC was 0.839- 0.944 for CRP. A CRP value ≥ 3.58 mg/L shows a sensitivity of 83.1% and a specificity of 72.3% in assessing MDD patients. These findings are consistent with other studies [9,15-19]. Diagnosis of Major Depressive Disorder (MDD) is done by clinical criteria. Tumor necrosis factor-α (TNF-α), interleukine-6 (IL-6), interleukine1β (IL-1β), CD4/CD8 ratio, haptoglobin, PGE2, corticotrophin releasing hormone are useful biomarkers. These tests are expensive and time consuming not easily available. Neutrophil/Lymphocyte Ratio (NLR), Red blood cell Distribution Width (RDW) Mean Platelet Volume (MPV) and C - Reactive Protein (CRP) have been suspected as the predictors of various diseases as well as MDD. NLR, RDW and MPV are derived from Complete Blood Count (CBC). CBC and CRP is less expensive and mostly available test all over the world as well as in Bangladesh. So, observations of this study were within international norms. Our data indicate that, RDW, NLR, MPV and CRP may play role in the assessment of MDD.



Table 9: Distribution of the study population according to groups with CRP (n=130).

  

    
CRP  (mg/L) 
    
Case    (n=65) 
    
Control    (n=65) 
    
p value*
  
  

    
 
    
n
    
%
    
n
    
%
    
 
  
  

    
Raised (>3.58)
    
54
    
83.1
    
18
    
27.7
    
0.001s
  
  

    
Normal
    
11
    
16.9
    
47
    
72.3
  
  

    
Total 
    
65
    
100
    
65
    
100
    
 
  

Table 9: Distribution of the study population according to groups with CRP
(n=130).

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Table 10: Validity of diagnostic tests.

  

    
Diagnostic tests 
    
Sensitivity 
    
Specificity 
    
PPV 
    
NPV 
    
Accuracy 
  
  

    
RDW CV (%)
    
73.8%
    
72.3%
    
72.7%
    
73.4%
    
73.1%
  
  

    
MPV (fl)
    
89.2%
    
81.5%
    
82.9%
    
88.3%
    
85.4%
  
  

    
NLR
    
84.6%
    
73.8%
    
76.4%
    
82.8%
    
79.2%
  
  

    
CRP (mg/L)
    
83.1%
    
72.3%
    
75.0%
    
81.0%
    
77.7%
  

Table 10: Validity of diagnostic tests.

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Conclusion

It is concluded that RDW, NLR, MPV and CRP were found higher in patients with MDD than healthy controls. These are less expensive and mostly available test. NLR, RDW-CV and MPV can detect during CBC count by auto-analyzer. Sensitivity and specificity are also more. So these parameters can be used as a valuable and effective tool for the management of MDD and treatment follow up.

References

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5. Firoz AHM, Karim ME, Alam MF. ‘Community based multi-centric service oriented research on mental illness with focus on awareness, prevalence, care, acceptance and follow-up in Bangladesh’. Manual on Mental Health for primary health care physicians; 2nd edn. NIMH & WHO. 2017.
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8. Zorrilla EP, Luborsky L, McKay JR, Rosenthal R, Houldin A, Tax A, et al. ‘The relationship of depression and stressors to immunological assays: a metaanalytic review’. Brain Behav Immun. 2001; 15: 199–226.
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Citation: Kanti Paul T, Paul D, Shamsun Nahar J, Ahmed S, Saiful Islam M, Ferdoushi S, et al. Value of Neutrophil (Lymphocyte Ratio), Red Blood Cell Distribution Width, Mean Platelet Volume and C - Reactive Protein in the Assessment of Major Depressive Disorder. Austin J Clin Pathol. 2017; 4(2): 1051.

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