Leukocyte function assessed via serial microlitre sampling of peripheral blood from sepsis patients correlates with disease severity

In this study, we demonstrate that serial assessment of leukocyte function from 50-μL of peripheral blood from sepsis patients provides significantly more prognostic information than leukocyte counting and is more highly correlated to disease severity.

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Sepsis is a life-threatening condition caused by a dysregulated immune response to uncontrolled infection and tissue injury [1]. Neutrophils and monocytes are among the first cells to respond for defending the host and play a critical role in controlling the infection, minimizing tissue injury, and promoting a return to homeostasis and health [2]. If these host innate immune responses are excessive and unregulated, additional and irreversible damage to host tissues can occur, resulting in organ dysfunction and death [2]. Using preclinical experimental mouse models of sepsis, interventions that regulate leukocyte function can decrease organ injury and lessen the severity of infection [3]. Therefore, the assessment of leukocyte activation and function may prove critical to the development of new host-directed therapies to restrain the over-exuberant inflammatory response and organ failure in human sepsis.

In patient care, currently available methods for diagnosis and point-of-care monitoring of sepsis are blunt.  Clinicians principally rely on clinical criteria and peripheral blood leukocyte counts.  Assessment of leukocyte function is not routinely determined. The correlation between peripheral blood leukocyte counts and sepsis diagnosis, sepsis severity, and most clinical outcomes is poor. Thus, developing new methods to diagnose and monitor sepsis is a research priority [4]. Bioengineering approaches to this clinical problem are beginning to bear fruit. Recently, measurement of patterns of neutrophil spontaneous motility in microfluidic chambers was found to correlate with the diagnosis of sepsis [5]. This exciting observation suggests that the assessment of leukocyte function rather than number alone has promise as a clinically relevant biomarker for the host immune response in sepsis.

To address this important unmet clinical need, the research groups of Drs. Baron, Han, Levy and Voldman from Brigham and Women’s Hospital and Massachusetts Institute of Technology collaborated to develop a new bioengineering approach for leukocyte sample preparation in microliter quantities of peripheral blood that enabled routine assessment of leukocyte function. Just published in Nature Biomedical Engineering [6], we applied a closed-loop inertial microfluidic platform for label free isolation of leukocytes from a small aliquot of peripheral blood (50 μl) (Fig. 1a) and assessed leukocyte phenotype and function from a cohort of 18 hospitalized patients with sepsis and 10 healthy subjects. We demonstrated using repeated sampling of sepsis patients over 7 days that quantification of leukocyte phenotype and function outperformed the complete blood count parameters (Fig. 1b,c) and were more highly correlated to sepsis severity (Fig. 1d,e). 

Sepsis is a devastating illness with limited diagnostic and therapeutic avenues. The findings presented in our study are a significant step forward in building a point-of-care diagnostic platform for monitoring the host immune response.  Increased precision in our routine assessment of immune responses will enable precision medicine approaches for monitoring the clinical course of critical illnesses, such as sepsis, and patient responses to therapeutic interventions.  

Figure 1. Sample Sparing Assessment of Leukocyte Function Correlates with Disease Severity. (a) A schematic of the closed-loop operation of the spiral microfluidics system. Two-dimensional score plots and loading plots from multivariate principal component analyses were determined for (b) routine clinical CBC parameters, (c) PMN functional responses (n = 9 healthy subjects and n = 11 sequential samples from 5 sepsis patients). The relationships between SOFA score (days 0–7) and (d) total PMNs (n = 32 separate biospecimens from 14 sepsis patients) and (e) ΔmedIDP (n = 23 from 13 sepsis patients) from sepsis patients at days 0, 3 and 7. The Pearson correlation r-value and significance are noted and regression lines are shown. Figure adapted from manuscript [6].


References:

1. Delano, M.J. and P.A. Ward, Sepsis-induced immune dysfunction: can immune therapies reduce mortality?J Clin Invest, 2016. 126(1): p. 23-31.

2. Serhan, C.N., Pro-resolving lipid mediators are leads for resolution physiology.Nature, 2014. 510(7503): p. 92-101.

3. Spite, M., et al., Resolvin D2 is a potent regulator of leukocytes and controls microbial sepsis.Nature, 2009. 461(7268): p. 1287-91.

4. Reddy, B., Jr., et al., Point-of-care sensors for the management of sepsis.Nat Biomed Eng, 2018. 2(9): p. 640-648.

5. Ellett, F., et al., Diagnosis of sepsis from a drop of blood by measurement of spontaneous neutrophil motility in a microfluidic assay.Nat Biomed Eng, 2018. 2(4): p. 207-214.

6. Jundi, B., et al., Leukocyte function assessed via serial microlitre sampling of peripheral blood from sepsis patients correlates with disease severity.Nat Biomed Eng, 2019. 3(12): p. 961-973.

 

 


Bruce D. Levy

Professor, Brigham and Women's Hospital/Harvard Medical School

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