Introduction
Materials and methods
Study population
RNA extraction, cDNA synthesis, and quantitative real-time PCR
Gene | Forward primer (5′ → 3′) | Reverse primer (5′ → 3′) |
---|---|---|
TLR1 | GAAGATTTCTTGCCACCCTAC | GAACACAATGTGCAGACTCTC |
TLR2 | CTGGACAATGCCACATAC | CTAATGTAGGTGATCCTG |
TLR4 | CAGAACTGCAGGTGCTGG | GTTCTCTAGAGATGCTAG |
TLR6 | CTATTGTTAAAAGCTTCCATTTTGT | ACCTGAAGCTCAGCGATGTAGTTC |
Β-Actin | ACTTAGTTGCGTTACACCCTT | GTCACCTTCACCGTTCCA |
Statistical analysis
Results
Demographics of the study subjects
Trait | COVID-19 subjects (N = 90) | Controls (N = 50) |
---|---|---|
Gender; Male/ Female (N, %) | 48 (53.3%)/ 42 (46.7%) | 28 (56%)/ 22 (44%) |
Smoker/ Non-smoker | 41 (45.5%)/ 49 (54.5%) | 21 (42%)/ 29 (58%) |
Age (Year, mean ± SD) | 49.2 ± 13.4 | 48.3 ± 12.6 |
Duration of COVID-19 (Day) | 10.9 ± 1.4 | – |
Oxygen saturation | 91.8 ± 7.1 | – |
Systolic BP (mmHg) | 138.7 ± 20.4 | – |
Diastolic BP (mmHg) | 75.8 ± 9.9 | – |
WBC (cells/mm3) | 9451 ± 1877.3 | – |
Neutrophil–lymphocyte ratio | 10.4 ± 6.9 | – |
ALP (IU/L) | 244.8 ± 42.9 | – |
AST (IU/L) | 35.7 ± 11.3 | – |
ALT (IU/L) | 36.4 ± 7.5 | – |
LDH (IU/L) | 408.2 ± 97.1 | – |
CRP (mg/L) | 5.1 ± 1.2 | – |
ESR (mm/h) | 22.7 ± 10.3 | – |
BMI (kg/m2) | 27.9 ± 7.1 | – |
Total cholesterol (mg/dl) | 223.2 ± 41.9 | – |
TG (mg/dl) | 173.7 ± 51.5 | – |
LDL (mg/dl) | 144.2 ± 35.6 | – |
HDL (mg/dl) | 49.2 ± 14.1 | – |
Creatinine (mg/dl) | 1.87 ± 0.49 | – |
BUN (mg/dl) | 24.4 ± 13.2 | – |
FBS (mg/dl) | 101.5 ± 32.7 | – |
D-dimer (ng/ml) | 1.74 ± 0.21 | – |
Cardiovascular diseases | 12 (13.3%) | – |
Diabetes | 11 (12.2%) | – |
Hypertension | 16 (17.8%) | – |
Fever | 60 (66.6%) | – |
Cough | 59 (65.5%) | – |
Dyspnea | 55 (61.1%) | – |
Sputum | 39 (43.3%) | – |
Vomiting/diarrhea | 33 (36.7%) | – |
Methylprednisolone use | 29 (32.2%) | – |
Remdesivir use | 21 (23.3%) | – |
Azithromycin use | 12 (13.3%) | – |
Anticoagulation therapy | 16 (17.8%) | – |
mRNA expression of TLRs
Item | TLR3 Fold change (P value) | TLR7 Fold change (P value) | TLR8 Fold change (P value) | TLR9 Fold change (P value) |
---|---|---|---|---|
COVID-19- Group A vs. Control-Group I | 3.4 (0.032) | 2.0 (0.041) | 2.8 (0.019) | 2.1 (0.016) |
COVID-19- Group A vs. Control-Group II | 2.9 (0.011) | 2.2 (0.020) | 2.9 (0.040) | 2.5 (0.001) |
COVID-19- Group A vs. Control-Group III | 2.2 (0.004) | 2.5 (0.038) | 3.1 (0.018) | 3.0 (0.026) |
COVID-19- Group B vs. Control-Group I | 2.1 (0.022) | 1.9 (0.025) | 2.0 (0.039) | 2.2 (0.024) |
COVID-19- Group B vs. Control-Group II | 2.3 (0.016) | 2.1 (0.033) | 2.3 (0.022) | 2.4 (0.019) |
COVID-19- Group B vs. Control-Group III | 2.4 (0.017) | 2.0 (0.009) | 2.2 (0.004) | 2.9 (0.010) |
COVID-19- Group C vs. Control-Group I | 1.2 (0.099) | 1.4 (0.18) | 1.1 (0.087) | 1.0 (0.891) |
COVID-19- Group C vs. Control-Group II | 1.2 (0.084) | 1.2 (0.082) | 1.0 (0.530) | 1.1 (0.787) |
COVID-19- Group C vs. Control-Group III | 1.3 (0.20) | 1.0 (0.139) | 1.2 (0.449) | 1.4 (0.094) |
Correlation of TLRs expression with clinicopathological findings
Item | TLR3 | TLR7 | TLR8 | TLR9 |
---|---|---|---|---|
Age | rho = 0.23, P = 0.098 | rho = 0.18, P = 0.254 | rho = 0.16, P = 0.650 | rho = 0.10, P = 0.309 |
Duration of COVID-19 | rho = 0.29, P = 0.088 | rho = 0.24, P = 0.074 | rho = 0.25, P = 0.147 | rho = 0.27, P = 0.205 |
Oxygen saturation | rho = 0.37, P = 0.041 | rho = 0.30, P = 0.026 | rho = 0.35, P = 0.022 | rho = 0.39, P = 0.028 |
Systolic BP | rho = 0.11, P = 0.410 | rho = 0.16, P = 0.201 | rho = 0.20, P = 0.085 | rho = 0.13, P = 0.416 |
Diastolic BP | rho = 0.11, P = 0.265 | rho = 0.21, P = 0.247 | rho = 0.13, P = 0.497 | rho = 0.20, P = 0.122 |
WBC | rho = 0.55, P = 0.020 | rho = 0.59, P = 0.029 | rho = 0.44, P = 0.012 | rho = 0.41, P = 0.034 |
Neutrophil–lymphocyte ratio | rho = 0.62, P = 0.012 | rho = 0.60, P = 0.013 | rho = 0.49, P = 0.011 | rho = 0.39, P = 0.036 |
ALP | rho = 0.16, P = 0.211 | rho = 0.10, P = 0.854 | rho = 0.19, P = 0.099 | rho = 0.18, P = 0.388 |
AST | rho = 0.18, P = 0.248 | rho = 0.15, P = 0.411 | rho = 0.10, P = 0.815 | rho = 0.13, P = 0.867 |
ALT | rho = 0.12, P = 0.381 | rho = 0.09, P = 0.880 | rho = 0.19, P = 0.231 | rho = 0.19, P = 0.299 |
LDH | rho = 0.39, P = 0.041 | rho = 0.41, P = 0.008 | rho = 0.49, P = 0.030 | rho = 0.25, P = 0.090 |
CRP | rho = 0.85, P = 0.001 | rho = 0.80, P = 0.004 | rho = 0.78, P = 0.010 | rho = 0.48, P = 0.035 |
ESR | rho = 0.71, P = 0.005 | rho = 0.62, P = 0.011 | rho = 0.65, P = 0.029 | rho = 0.40, P = 0.044 |
BMI | rho = 0.13, P = 0.655 | rho = 0.11, P = 0.249 | rho = 0.08, P = 0.709 | rho = 0.16, P = 0.380 |
Total cholesterol | rho = 0.20, P = 0.568 | rho = 0.09, P = 0.744 | rho = 0.20, P = 0.516 | rho = 0.16, P = 0.246 |
TG | rho = 0.13, P = 0.214 | rho = 0.19, P = 0.501 | rho = 0.25, P = 0.077 | rho = 0.16, P = 0.504 |
LDL | rho = 0.09, P = 0.209 | rho = 0.19, P = 0.640 | rho = 0.15, P = 0.309 | rho = 0.14, P = 0.506 |
HDL | rho = 0.11, P = 0.246 | rho = 0.16, P = 0.407 | rho = 0.15, P = 0.230 | rho = 0.16, P = 0.488 |
Creatinine | rho = 0.26, P = 0.087 | rho = 0.11, P = 0.501 | rho = 0.13, P = 0.249 | rho = 0.18, P = 0.202 |
BUN | rho = 0.11, P = 0.480 | rho = 0.23, P = 0.509 | rho = 0.11, P = 0.600 | rho = 0.14, P = 0.407 |
FBS | rho = 0.13, P = 0.260 | rho = 0.18,
P = 0.354 | rho = 0.08, P = 0.580 | rho = 0.18, P = 0.404 |
D-dimer | rho = 0.08, P = 0.333 | rho = 0.16, P = 0.240 | rho = 0.11, P = 0.266 | rho = 0.25, P = 0.088 |