Abstract
Objective
We aimed to investigate whether there is a relationship between diaphragm thickness and disease severity in Covid-19 pregnant subgroups.
Material and methods
In this prospective study 100 pregnant patients were enrolled. Thickness of the diaphragm muscle at end-expiration was measured using B-Mode US. Hemoglobin,WBC, NLR, procalcitonin and LDH levels were measured.
Results
There was a statistically significant difference between the groups in terms of diaphragm thickness, and the diaphragm thickness was thinner in the severe disease group (p < 0.001). There was no statistically significant difference between the groups with mild to moderate disease severity (p = 0.708).
Conclusion
Covid-19 patients who developed serious infection has thinner diaphragms than those who did not. Low diaphragm muscle thickness at the outset of Covid-19 disease, may predispose to poor clinical outcomes. Diaphragmatic ultrasound may be a promising tool to evaluate the risk of Covid-19 disease severity.
Keywords: Pregnancy, Ultrasound, Covid-19, Diaphragmatic thickness
Introduction
The Covid-19 pandemic caused by the SARS-CoV-2 infection, which was detected in Wuhan, China in December 2019 and spread rapidly all over the world, infected more than 207 million people and killed more than 4 million people. Although most cases are characterized by minimal flulike constitutional symptoms or may even be asymptomatic, some patients develop a severe pneumonia that may lead to acute respiratory distress syndrome, multiorgan failure, and death [1]. With disease progression, patients may become severely dyspneic and unable to maintain adequate ventilation by movement of their inspiratory muscles. Systemic hyperinflammation and procoagulation play an important role in the pathophysiology of these severe forms [2]. In particular, catabolism of muscle proteins has been associated with muscle weakness and/or atrophy, increased morbidity and mortality [3].
The diaphragm is the main muscle of respiration. A wide variety of factors such as sepsis, medications, and multiorgan dysfunction syndrome can cause acute or chronic diaphragmatic weakness in critically ill patients [4]. Numerous studies have shown that the change in respiratory and limb muscle size and quality predicts prolonged ventilator dependence and poor clinical outcomes in patients infected Covid-19 [[5], [6], [7], [8]].
However, the acute effect of severe Covid-19 in muscle loss and functional impairment is unknown. The use of ultrasonography, an easily available and non-invasive technique for measuring diaphragm thickness, is increasingly being used to evaluate changes in both muscle size and quality over time.
In our study, diaphragmatic muscle thickness is measured in pregnant Covid-19 infected patients with B-Mode US to determine the relationship between thickness and disease severity. We aimed to investigate whether there is a relationship between diaphragm thickness and disease severity in Covid-19 pregnant subgroups.
Materials and methods
This prospective study was performed between February 2021 to June 2021. Ethical approval was obtained from the Ministry of Health Ankara City Hospital, Ethical Review Board (No: E2-21-75). The present study was conducted following the principles of the Declaration of Helsinki. All patients were extensively informed of study design. Written informed consent was obtained by all patients enrolled. Pregnant patients (n:100) who were hospitalized for Covid-19 were scanned as a part of their routine care.
Patients who had COVID-19 infection and were hospitalized and followed up, were included in the study. Those who had neuromuscular diseases, maternal systemic diseases (diabetes, cardiovascular diseases, hypertensive diseases of pregnancy, asthma, thromboembolic disorders, inflammatory bowel diseases, autoimmune connective tissue diseases, other infectious diseases) and multifetal pregnancy were excluded. None of the patients needed intensive care and mechanical ventilation.
Groups were divided into 3 as mild, moderate and severe. The severity of COVID-19 was determined by the need for inhaler oxygen therapy as mild (a respiratory rate of 30 breaths per minute, blood O2 saturation of <93% on room air and need of oxygen under 2 L/min), moderate (a respiratory rate of 30 breaths per minute, blood O2 saturation of <93% on room air and need of oxygen 2–5 L/min) or severe (a respiratory rate of 30 breaths per minute, blood O2 saturation of <93% on room air and need of oxygen more than 5 L/min). The patient characteristics of the study population are shown in Table 1 .
Table 1.
Demographic and clinical characteristics of cases regarding for severity of disease.
| Mild (n = 36) | Moderate (n = 40) | Severe (n = 24) | p-value | |
|---|---|---|---|---|
| Age (years) | 29.0 ± 5.9 | 30.0 ± 5.6 | 30.6 ± 4.5 | 0.717a |
| BMI (kg/m2) | 27.7 ± 2.5 | 28.5 ± 2.9 | 28.3 ± 2.7 | 0.626a |
| Gravidity | 2 (1–10) | 2 (1–7) | 2 (1–4) | 0.934b |
| Gestational age (weeks) | 36.5 (30–39) | 38 (31–39) | 34.5 (31–38) | 0.107b |
Data were displayed as mean ± SD or median (min–max); where applicable.
One-Way ANOVA.
Kruskal Wallis test.
Thickness of the diaphragm muscle at end-expiration was measured using B-Mode US. All patients underwent B-mode US examinations in supine position. The diaphragm muscle was assessed on a portable ultrasound system (Logiq E, GE). All subjects underwent B-mode ultrasound examination. The diaphragm was visualized by placing the probe in the ninth intercostal space perpendicular to two ribs. The diaphragm muscle is located between two parallel echoic lines which named as the diaphragmatic pleura and the peritoneal membrane. Thickness was measured by placing the calipers inside the hyperechoic lines. Three diaphragm thickness measurement of the right diaphragm were recorded at end-expiration and averaged (Fig. 1 ). The patients were evaluated by the same radiologist who had no knowledge of the groups. In order to prevent the study from being affected by the treatment methods and differences, ultrasonographic diaphragmatic thickness measurement of all patients was performed on the day of hospitalization before the treatment was started.
Fig. 1.
Ultrasonographic imaging of measurement.
Blood samples were obtained before any medication was taken. The hemoglobin, WBC, NLR, procalcitonin and LDH levels among the groups were measured for all patients in routine monitoring.
All patients were managed according to the national guidelines at the time of the study by a multidisciplinary team comprising obstetricians, radiologists and infectious disease specialists [9].
Statistical analysis was performed using IBM SPSS Statistics ver. 25.0 software (IBM Corporation, Armonk, NY, US). Whether the assumptions of normal distribution and, variance homogeneity were met or not were investigated by Shapiro–Wilk and, Levene tests; respectively. Data were expressed as mean ± SD, median (min–max) or median [25th–75th] percentiles; where applicable. While the mean differences among groups were compared One-Way ANOVA, otherwise the Kruskal Wallis test was applied for the comparison of quantitative data which parametrical test assumptions were not met. When the p-values from One-Way ANOVA or Kruskal Wallis tests were statistically significant, post-hoc Tukey's HSD or Dunn–Bonferroni test were used to determine which group differ from which others. The optimal threshold for diaphragm thickness in order to predict the severity of disease was evaluated ROC analysis as giving the maximum sum of sensitivity and specificity for the significant test. Sensitivity, specificity, positive and negative predicted values and accuracy for diaphragm thickness were also calculated. A p value less than 0.05 was considered statistically significant.
Results
Over the study period, we treated 100 pregnant Covid-19 infected patients in the inpatient service. Their clinical characteristics, laboratory data and imaging findings at admission are summarized in Table 1. There was no statistically significant difference between the groups with mild, moderate and severe disease severity in terms of mean age, body mass index, gravida and gestational weeks (p > 0.05).
Table 2 shows the comparisons made between the groups in terms of biochemical measurements. There was no statistically significant difference in terms of hemoglobin, WBC, NLR, procalcitonin and LDH levels among the groups with mild, moderate and severe disease severity, respectively (p > 0.05).
Table 2.
The comparisons among study groups in terms of laboratory measurements.
| Mild (n = 36) | Moderate (n = 40) | Severe (n = 24) | p-value | |
|---|---|---|---|---|
| Hemoglobin | 11.6 ± 1.67 | 11.6 ± 1.03 | 11.5 ± 1.28 | 0.966a |
| WBC | 8.8 ± 2.7 | 8.8 ± 3.1 | 6.8 ± 3.1 | 0.132a |
| NLR | 4.4 [3.7–5.5] | 4.8 [3.9–6.4] | 5.5 [4.3–9.0] | 0.138b |
| CRP | 16.5 [7.2–39.7]c | 9.0 [2.5–23.0]d | 47.5 [36.2–64.5]c,d | 0.004b |
| Ferritin | 13.5 [5.0–35.7]c | 17.5 [11.0–29.0] | 53.5 [18.0–313.7]c | 0.029b |
| d-Dimer | 1.6 [1.1–2.3]c | 2.3 [1.6–3.4] | 2.5 [1.7–4.2]c | 0.026b |
| Procalcitonin | 0.03 [0.03–0.08] | 0.05 [0.03–0.07] | 0.08 [0.03–0.17] | 0.127b |
| LDH | 221.5 [207.0–302.7] | 240.0 [219.0–274.5] | 261.0 [213.5–439.0] | 0.329b |
Data were displayed as mean ± SD or median [25th–75th] percentiles; where applicable.
One-Way ANOVA.
Kruskal Wallis test.
Mild vs Severe (p < 0.05),
Moderate vs Severe (p = 0.004).
There was a statistically significant difference between the groups in terms of CRP levels (p = 0.004). The situation that caused this difference; the CRP level of the group with severe disease severity was higher than the mild and moderate groups (p = 0.041 ve p = 0.004). There was no statistically significant difference between the groups with mild to moderate disease severity (p > 0.999).
There was a statistically significant difference between the groups in terms of ferritin levels (p = 0.029). The ferritin level of the group with severe disease severity was higher than the mild group (p = 0.025). Ferritin levels were statistically similar between mild and moderate disease severity groups and between moderate and severe disease severity groups (p > 0.999 and p = 0.165).
There was also a statistically significant difference between the groups in terms of d-Dimer levels (p = 0.026). d-Dimer level was higher in the severe disease group than in the mild disease group (p = 0.039). d-Dimer levels were statistically similar between mild and moderate disease severity groups and between moderate and severe disease severity groups (p = 0.106 and p > 0.999).
In Table 3 , it was evaluated whether there was a statistically significant change in the diaphragm thickness according to the severity of the disease. There was a statistically significant difference between the groups in terms of diaphragm thickness, and the diaphragm thickness was thinner in the severe disease group (p < 0.001). There was no statistically significant difference between the groups with mild to moderate disease severity (p = 0.708) (Fig. 2 ). Management of patients with infected Covid-19 are shown in Table 4 .
Table 3.
The thickness of diaphragm according to the severity of disease.
| N | Thickness of diaphragm | |
|---|---|---|
| Mild | 36 | 3.49 ± 0.39a |
| Moderate | 40 | 3.39 ± 0.41b |
| Severe | 24 | 2.70 ± 0.30a,b |
| p-valuec | <0.001 |
Data were shown as mean ± SD.
Mild vs Severe (p < 0.001),
Moderate vs Severe (p < 0.001).
One-Way ANOVA.
Fig. 2.
Thickness of diaphragma.
Table 4.
Management of pregnant women with Covid-19.
| COVID-19 therapy | Mild (n = 36) | Moderate (n = 40) | Severe (n = 24) |
|---|---|---|---|
| Low-molecular weight heparin (n, %) | 36 (100%) | 40 (100%) | 24 (100%) |
| Hydroxychloroquine (n, %) | – | 7 (17,5%) | 19 (79,1%) |
| Systemic corticosteroid (n, %) | – | 2 (5%) | 24 (100%) |
| Favipiravir (n, %) | – | – | 5 (20,8%) |
| Lopinavir-ritonavir (n, %) | – | – | 3 (12,5%) |
| N-acetylcysteine (n, %) | – | – | 3 (12,5%) |
| Antibiotherapy for other pathogens (n, %) | 36 (100%) | 40 (100%) | 24 (100%) |
Note: All treatments were started after diaphragmatic thickness measurement to avoid muscle mass changes.
Finally, It was investigated whether diaphragm thickness was a statistically significant predictor of disease severity by ROC analysis. The area under the ROC curve of the diaphragm thickness was found to be statistically significant in order to differentiate the group with severe disease from those with mild or moderate disease (AUC = 0.925, 95% CI: 0.840–1.000, p < 0.001) (Fig. 3 ). The best cut-off point of the diaphragm thickness in distinguishing the group with severe disease and the groups with mild or moderate severity is 2.95 mm, the sensitivity of the diaphragm thickness at this point is 91.7%, the selectivity 89.5%, positive and negative predictive values, respectively; 73.3% and 97.1%, with a diagnostic accuracy of 90.0%.
Fig. 3.
The best cut-off point of the diaphragm thickness in distinguishing serious disease is 2.95 mm, the sensitivity of the diaphragm thickness at this point is 91.7%, the selectivity is 89.5%.
Discussion
To the best of our knowledge, this is the first study to investigate the relationship between the severity of Covid-19 disease and diaphragm thickness in pregnant patients. The main findings of this prospective study are that; firstly patients who developed severe covid 19 disease had thinner diaphragm than those who did not and secondly CRP, ferritin and D dimer levels were independent significant predictors of severe Covid-19 disease.
Diaphragm thickness measurements in Covid-19 patients has been investigated in few studies in the literature. Corradi et al. measured diaphragmatic thickness in patients with laboratory-confirmed Covid-19 infection. They found more adverse outcomes in patients with thinner diaphragms. They thought that this situation might develop secondary to muscle loss caused by exaggerated proteolysis due to Covid-19 infection [10].
Sonographic assessment of the diaphragm thickness was recently evaluated for disease progression in Covid-19 patients. Umbrello et al. compared that the size of the diaphragm muscle between Covid-19 survivors and non-survivors. Muscle ultrasound was performed in all thirty-six patients enrolled in their study. They found that diaphragm sizes were significantly reduced during the course of the disease and this reduction was significantly higher in non-survivors [11].
Sklar et al. shown that decreases of diaphragm thickness during the early course of mechanical ventilation in Covid-19 patients associated with a poor clinical outcome in their study. Similarly, we speculated that diaphragm thickness might be at significant risk factor for Covid-19 in our study. To do so, we used muscle ultrasound which is a non-invasive and cheap imaging modality for muscle assessment [12].
It has been shown in previous studies that sepsis causes a myopathy of the respiratory muscles. Reduction in existing muscle thickness is a major cause of mortality and long-term morbidity in patients [13]. Similarly we found that the results of our study were also the diaphragm thickness of the group with severe disease course and the groups with mild or moderate severity were different, and it was thinner in the severe course. Measuring diaphragm muscle thickness in advance may be useful in predicting the course of the disease.
Although the mechanical effects of Covid-19 on the muscles are not fully understood due to the lack of sufficient studies; in mouse models of SARS-CoV-1, a rapid reduction of 20% in body mass was found 4 days after infection. SARS-CoV-1 infection caused muscle fiber necrosis, immune cell infiltration, and diffuse muscle fiber atrophy [14]. In addition to direct viral infection, cytokines and released proinflammatory molecules can lead to pathological changes in muscle tissue. Similarly proinflammatory molecules triggered by Covid-19 can accelerate muscle fiber proteolysis and decrease protein synthesis. C-reactive protein (CRP), which is used as a common biomarker in the determination of general inflammation, has been found to be much higher in patients with severe Covid-19 compared to healthy controls [15]. Similarly, we found CRP levels higher in the severe disease group in our study.
Ferritin levels measurement has diagnostic value for Covid-19. Ferritin levels are too high due to secondary hemophagocytic lymphohistiocytosis and cytokine storm syndrome seen in severe Covid-19 patients and are indicative of poor prognosis [16]. There is also a correlation between abnormal coagulation parameters and adverse outcome. Significantly higher plasma D-dimer levels were measured in patients who died from Covid-19 infection [17]. Clinical laboratory data are indisputably important in the diagnosis of Covid-19 disease and in the evaluation of the severity of the disease. Decreased levels of ferritin and D-dimer are the most commonly reported laboratory findings for COVID-19 in the literature [[18], [19], [20]]. We think that additional evaluation of diaphragm thickness may be useful to distinguish between severe and mild Covid-19 infection.
This study has several limitations. First, it was conducted in a small number of Covid-19 infected pregnant patients. Second, the observational nature and limited sample size of the present study do not allow for generalizations. Therefore, our results should be considered as suggestive for larger studies.
In conclusion Covid-19 patients who developed serious infection has thinner diaphragms than those who did not. Low diaphragm muscle thickness at the outset of Covid-19 disease, may predispose to poor clinical outcomes. It is possible that the severity of the infection play a crucial role in the observed loss of diaphragmatic muscle mass or early changes in muscle size and may potentially be related to the severity of the disease. Ultrasound that is used in our routine clinical practice is a well-tolerated noninvasive and inexpensive imaging modality. It can be preferred especially because it is a routine part of the examination of pregnant patients and easily applied to both inpatients and outpatients. Diaphragmatic ultrasound may be a promising tool to evaluate the risk of Covid-19 disease severity.
Ethical approval
All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.
Author contributions
Conceived and designed the experiments: EUO, GNB, NO, OMT. Performed the experiments: EUO, GNB, HLK, NO, OMT. Analyzed the data: EUO, GNB, DA, BE, GY. Wrote the paper: EUO, GNB.
Financial or funding support
Authors received no funding for completion of the study or manuscript.
Declaration of competing interest
All the authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
None declared.
References
- 1.Grasselli G., Zangrillo A., Zanella A., Antonelli M., Cabrini L., Castelli A., et al. Baseline characteristics and outcomes of 1591 patients infected with SARS-CoV-2 admitted to ICUs of the lombardy region. Italy. Jama. 2020;323(16):1574–1581. doi: 10.1001/jama.2020.5394. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 2.Qin C., Zhou L., Hu Z., Zhang S., Yang S., Tao Y., et al. Dysregulation of immune response in patients with coronavirus 2019 (COVID-19) in wuhan, China. Clin Infect Dis. 2020;71(15):762–768. doi: 10.1093/cid/ciaa248. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 3.Gruther W., Benesch T., Zorn C., Paternostro-Sluga T., Quittan M., Fialka-Moser V., et al. Muscle wasting in intensive care patients: ultrasound observation of the M. quadriceps femoris muscle layer. J Rehabil Med. 2008;40(3):185–189. doi: 10.2340/16501977-0139. [DOI] [PubMed] [Google Scholar]
- 4.Demoule A., Jung B., Prodanovic H., Molinari N., Chanques G., Coirault C., et al. Diaphragm dysfunction on admission to the intensive care unit. Prevalence, risk factors, and prognostic impact-a prospective study. Am J Respir Crit Care Med. 2013;188(2):213–219. doi: 10.1164/rccm.201209-1668OC. [DOI] [PubMed] [Google Scholar]
- 5.Demoule A., Molinari N., Jung B., Prodanovic H., Chanques G., Matecki S., et al. Patterns of diaphragm function in critically ill patients receiving prolonged mechanical ventilation: a prospective longitudinal study. Ann Intensive Care. 2016;6(1):75. doi: 10.1186/s13613-016-0179-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 6.Dres M., Dubé B.P., Mayaux J., Delemazure J., Reuter D., Brochard L., et al. Coexistence and impact of limb muscle and diaphragm weakness at time of liberation from mechanical ventilation in medical intensive care unit patients. Am J Respir Crit Care Med. 2017;195(1):57–66. doi: 10.1164/rccm.201602-0367OC. [DOI] [PubMed] [Google Scholar]
- 7.Goligher E.C., Dres M., Fan E., Rubenfeld G.D., Scales D.C., Herridge M.S., et al. Mechanical ventilation-induced diaphragm atrophy strongly impacts clinical outcomes. Am J Respir Crit Care Med. 2018;197(2):204–213. doi: 10.1164/rccm.201703-0536OC. [DOI] [PubMed] [Google Scholar]
- 8.Jung B., Moury P.H., Mahul M., de Jong A., Galia F., Prades A., et al. Diaphragmatic dysfunction in patients with ICU-acquired weakness and its impact on extubation failure. Intensive Care Med. 2016;42(5):853–861. doi: 10.1007/s00134-015-4125-2. [DOI] [PubMed] [Google Scholar]
- 9.Turkish Ministry of Health GDoPH. COVİD-19 (SARS-CoV-2 infection) guideline, scientific committee report.
- 10.Corradi F., Isirdi A., Malacarne P., Santori G., Barbieri G., Romei C., et al. Low diaphragm muscle mass predicts adverse outcome in patients hospitalized for COVID-19 pneumonia: an exploratory pilot study. Minerva Anestesiol. 2021;87(4):432–438. doi: 10.23736/S0375-9393.21.15129-6. [DOI] [PubMed] [Google Scholar]
- 11.Umbrello M., Guglielmetti L., Formenti P., Antonucci E., Cereghini S., Filardo C., et al. Qualitative and quantitative muscle ultrasound changes in patients with COVID-19-related ARDS. Nutrition. 2021;91–92 doi: 10.1016/j.nut.2021.111449. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 12.Sklar M.C., Dres M., Fan E., Rubenfeld G.D., Scales D.C., Herridge M.S., et al. Association of low baseline diaphragm muscle mass with prolonged mechanical ventilation and mortality among critically ill adults. JAMA Netw Open. 2020;3(2) doi: 10.1001/jamanetworkopen.2019.21520. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 13.Callahan L.A., Supinski G.S. Sepsis-induced myopathy. Crit Care Med. 2009;37(10 Suppl):S354–S367. doi: 10.1097/CCM.0b013e3181b6e439. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 14.McCray P.B., Jr., Pewe L., Wohlford-Lenane C., Hickey M., Manzel L., Shi L., et al. Lethal infection of K18-hACE2 mice infected with severe acute respiratory syndrome coronavirus. J Virol. 2007;81(2):813–821. doi: 10.1128/JVI.02012-06. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 15.Mao L., Jin H., Wang M., Hu Y., Chen S., He Q., et al. Neurologic manifestations of hospitalized patients with coronavirus disease 2019 in wuhan, China. JAMA Neurol. 2020;77(6):683–690. doi: 10.1001/jamaneurol.2020.1127. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 16.Mehta P., McAuley D.F., Brown M., Sanchez E., Tattersall R.S., Manson J.J. COVID-19: consider cytokine storm syndromes and immunosuppression. Lancet. 2020;395(10229):1033–1034. doi: 10.1016/S0140-6736(20)30628-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 17.Tang N., Li D., Wang X., Sun Z. Abnormal coagulation parameters are associated with poor prognosis in patients with novel coronavirus pneumonia. J Thromb Haemostasis. 2020;18(4):844–847. doi: 10.1111/jth.14768. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 18.Kaftan A.N., Hussain M.K., Algenabi A.A., Naser F.H., Enaya M.A. Predictive value of C-reactive protein, lactate dehydrogenase, ferritin and D-dimer levels in diagnosing COVID-19 patients: a retrospective study. Acta Inf Med. 2021;29(1):45–50. doi: 10.5455/aim.2021.29.45-50. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 19.Rostami M., Mansouritorghabeh H. D-dimer level in COVID-19 infection: a systematic review. Expet Rev Hematol. 2020;13(11):1265–1275. doi: 10.1080/17474086.2020.1831383. [DOI] [PubMed] [Google Scholar]
- 20.Cheng L., Li H., Li L., Liu C., Yan S., Chen H., et al. Ferritin in the coronavirus disease 2019 (COVID-19): a systematic review and meta-analysis. J Clin Lab Anal. 2020;34(10) doi: 10.1002/jcla.23618. [DOI] [PMC free article] [PubMed] [Google Scholar]