Advancing cell-based therapy in sepsis: An anesthesia outlook

Abstract

Sepsis poses a health challenge globally owing to markedly high rates of morbidity and mortality. Despite employing bundle therapy over two decades, approaches including transient organ supportive therapy and clinical trials focusing on signaling pathways have failed in effectively reversing multiple organ failure in patients with sepsis. Prompt and appropriate perioperative management for surgical patients with concurrent sepsis is urgent. Consequently, innovative therapies focusing on remedying organ injuries are necessitated. Cell therapy has emerged as a promising therapeutic avenue for repairing local damage to vital organs and restoring homeostasis during perioperative treatment for sepsis. Given the pivotal role of immune cell responses in the pathogenesis of sepsis, stem cell-based interventions that primarily modulate immune responses by interacting with multiple immune cells have progressed into clinical trials. The strides made in single-cell sequencing and gene-editing technologies have advanced the understanding of disease-specific immune responses in sepsis. Chimeric antigen receptor (CAR)-immune cell therapy offers an intriguing option for the treatment of sepsis. This review provides a concise overview of immune cell therapy, its current status, and the strides made in the context of sepsis research, discussing potential strategies for the management of patients with sepsis during perioperative stages.

Introduction

Based on the definition formulated in 2016, sepsis is a dysregulated host response resulting in life-threatening organ dysfunction [Figure 1]. It rapidly progresses to septic shock, concomitant with profound circulatory, cellular, and metabolic abnormalities. Thus, sepsis has a high mortality rate.^[1] It is estimated that sepsis affects approximately 49 million individuals annually, resulting in 11 million deaths worldwide.^[2] Sepsis and septic shock remain the major causes of death and impose a global burden owing to their increasing prevalence. To reduce the global burden of sepsis, the World Health Organization (WHO) has proposed the inclusion of sepsis as a priority research area to facilitate the development of innovative approaches to preventing, diagnosing, and treating sepsis.^[3]

Figure 1.

The third international consensus definitions for sepsis and septic shock (sepsis-3). The current definition for Sepsis-3 is as follows: sepsis is defined as a life-threatening organ dysfunction caused by a dysregulated host response to an infection. Sepsis-3 addresses the limitations of previous definitions that focused excessively on inflammation. The report redefined septic shock as a subset of sepsis in which particularly profound circulatory, cellular, and metabolic abnormalities form a continuum from severe sepsis to septic shock. The report reveals the inadequate specificity and sensitivity of the criteria for SIRS. SIRS: Systemic inflammatory response syndrome.

Medical Emergencies and Time-essential Syndrome Constituting by Sepsis and Septic Shock

It is estimated that 312.9 million major surgeries are performed globally per year.^[4] Sepsis ranks as the third most common perioperative comorbidity among surgical patients, with an incidence of 5.4%.^[5] It is a challenging condition to treat and a definitive cure is lacking. Severe trauma and major surgical procedures, caused typically by suppurative cholangitis, suppurative peritonitis, severe acute pancreatitis, and urinary tract infection perioperatively, are common precursors of sepsis. Chronic progression of a focal abscess or infection in the deep tissue or viscera can lead to sepsis, eventually requiring surgical intervention. In clinical settings, these patients require surgical interventions for infection control, debris removal, and drainage of purulent cavities [Figure 2].

Figure 2.

Removal of infected foci and repair of injured organs during perioperative sepsis. Sepsis induced by trauma and major surgical operations usually requires surgical interventions to remove necrotic tissue and infected foci. For these patients, surgical procedures, anesthesia and ventilation, and invasive procedures result in a massive release of DAMPs, implicated in aggravating the injured tissue and organs. The core treatment options for surgical septic cases are the removal of infected foci and repair of injured organs to maintain tissue perfusion and promote organ function recovery. DAMPs: Damage-associated molecular patterns.

However, these surgical procedures, along with agents to induce anesthesia and other invasive manipulations, can exacerbate tissue and organ damage. Under these circumstances, prompt and appropriate perioperative management of surgical patients with concurrent sepsis is critical. Anesthesiologists should be vigilant in monitoring, supporting potential organ dysfunction, and addressing tissue injuries.

Over the past three decades, global experts have put in significant efforts to enhance the clinical prognosis of septic shock and patients with sepsis. Since the inception of the Surviving Sepsis Campaign (SSC) in 2002, bundle treatment has been the primary approach to improve clinical outcomes in patients with sepsis.^[6] This protocol-driven therapy primarily includes early fluid resuscitation, prompt administration of antibiotics, hemodynamic stabilization, and organ-supportive care, aiming at curtailing the progression of organ dysfunction.^[6] Despite the implementation of organ-supportive strategies such as extracorporeal membrane oxygenation, continuous renal replacement therapy, artificial heart, and other options, mortality rates persistently remain high. The above-mentioned interventions for sepsis are all supportive, rather than targeted toward a specific pathophysiology.

Clinical patients with sepsis often succumb to multi-organ failure, stemming from the intricate interplay of enhanced inflammation and immunosuppression. The pathophysiology of patients with sepsis is complicated by different intensities of surgical trauma and the use of anesthetic drugs. However, epidemiological studies have identified the abdomen and lungs as the most common sites of infection. Gram-negative bacteria are predominant among all isolated microbes. From the perioperative perspective, surgical sepsis is heterogeneous. Notably, patients with preoperative sepsis or those worsening into intraoperative sepsis state always show compromised physical condition, especially an inherently unstable cardiovascular state during the operation.^[7] The majority of anesthetics including sedative drugs and opioid analgesics directly affect myocardial depression and vessel relaxation, further aggravating the reduction in preload and afterload. Together, this results in severe hypotension in septic patients. The risk for septic patients is high during the administration of anesthesia. Source control of infection determines the outcome in surgical patients with sepsis but once not completely cleared, it can lead to persistent infection and progression to unresolved septic shock and eventually, death. Furthermore, surgical intervention may allow the spread of infective materials into the bloodstream, thus worsening the patients’ hemodynamics and even requiring resuscitation. Appropriate and customized perioperative management of septic patients by anesthesiologists is particularly important.

Past clinical trials targeting single pathways or molecules against sepsis have been unsuccessful. However, with advancements in single-cell sequencing and gene-editing technology, delving into the intricate web of disease-specific immune responses that play a pivotal role in the pathogenesis of sepsis is possible [Figure 3].^[8,9,10] Particularly perioperatively, cell therapies tailored to specific sub-populations hold significant promise for clinical application. For example, macrophages are crucial cells within the innate immune system. These are present in diverse locations including the peripheral blood and vital target organs such as the lungs, liver, brain, and heart.^[11] Various sub-populations of macrophages assume distinct roles in different types, courses, and contexts of sepsis. For instance, tissue-resident macrophages (TRMs) are characterized by tissue-specific and niche-specific functions, such as the clearance of cellular debris, phagocytosis, tissue repair, and ultimately, the maintenance of homeostasis.^[11] The profound and dysregulated activation of macrophage sub-groups during sepsis can directly affect the outcome in these patients. From an anesthesia perspective, expeditious healing of injured organs is imperative to sustain tissue perfusion and facilitate organ function recovery. Intraoperative management based on a better understanding of the pathophysiology of sepsis is warranted. This review traces the progress of therapeutic strategies for sepsis, with a special focus on cell-based immunotherapy, and aims to offer fresh insights for clinical application.

Figure 3.

scRNA-seq for sepsis. ScRNA-seq initially involves the dissociation of tissues into single cells. Single cells are loaded onto a microfluidic chip and attached with a unique oligonucleotide sequence following cell identification. RNA from each single cell is reverse transcribed to obtain a pooled cDNA library. ScRNA-seq can reveal the heterogeneity and evolutionary trajectory of immune cells and contribute to the identification of immune cell subsets, thus providing new insights into disease-specific immune responses in the pathogenesis of sepsis and offering a theoretical basis for developing cell-based therapies for sepsis. DC: Dendritic cell; NK: Natural killer; scRNA-seq: Single-cell RNA sequencing.

Immune System Is the First Line of Defense Against Sepsis

Immune responses encompass two primary components—innate and adaptive immunity. Acute inflammation is primarily associated with innate immunity, which involves key cell types such as neutrophils and macrophages. Adaptive immunity involves T cells, B cells, and dendritic cells (DCs), and is responsible for immunosuppression and protection against secondary infections.^[12] The innate immune system is the first line of defense against pathogens invading the body.^[12] Upon encountering a threat, immunogenic foreign pathogen-associated molecular patterns (PAMPs) are promptly recognized by pattern recognition receptors (PRRs).^[13] This recognition is geared toward eradicating the invading microorganism and restoring homeostasis. PRRs can identify host molecules with damage-associated molecular patterns (DAMPs), which are released during periods of inflammatory stress (e.g., surgery, trauma, and tissue stress).^[13] DAMPs from damaged tissue or cells, along with PAMPs, are recognized by specific PRRs, setting off a cascade of events that lead to a systemically uncontrolled over-inflammation, referred to as a “cytokine storm”.^[14,15]

Exposure to anesthetic agents and major surgeries impair the immune response, which increases perioperative morbidity and mortality rates in patients with severe infection and sepsis. In the early stages of sepsis, the host’s immune responses are inundated with overwhelming inflammation, whereby a multitude of inflammatory mediators inflict damage to tissues and immune cells.^[12] In septic patients undergoing surgical procedures, expression of perioperative cytokines differs based on the intensities of surgical trauma and anesthetic agents, further affecting postoperative organ dysfunction. Dysregulation of cytokine expression including excess production of proinflammatory cytokines can induce hemodynamic instability and metabolic disorders. Theoretically, modulating these cytokines is a promising strategy for mitigating the drawbacks of sepsis-related tissue/organ dysfunction. Clinical trials attempting to neutralize inflammation and inflammatory mediators during sepsis have not demonstrated considerable improvements in overall survival or other indicators of good clinical prognosis.^[16,17,18] Toll-like receptors (TLRs), the main sub-family of PRRs, are crucial receptors associated with innate immunity. Specifically, TLR4 is the most attractive target for treating sepsis owing to its extensive participation in the activation of cellular and inflammatory responses during sepsis.^[14,19] A TLR4-specific inhibitor, TAK-242, prevents organ injuries in animal models of sepsis, but in published clinical trials, it failed to suppress inflammatory responses in patients with sepsis or septic shock.^[20] The large phase III multicenter ACCESS randomized trial did not reveal the benefit of treatment with eritoran, the TLR4/myeloid differentiation factor-2 (MD2) antagonist, in septic patients.^[21]

Among the reasons for the failure of anti-inflammatory strategies in patients with sepsis, one possible factor is the change in the syndrome over time.^[8] In both animal models and patients with sepsis, a systemic elevation in pro-inflammatory cytokine levels is neither consistently nor persistently observed.^[22] Conversely, anti-inflammatory cytokines, including interleukin-4 (IL-4), IL-10, and transforming growth factor beta (TGF-β), are consistently detected over extended periods.^[22] Anti-inflammatory immunological events develop concurrently or subsequently during sepsis. Subsequently, the advanced stage of sepsis is characterized by a profound state of immunosuppression.^[8,23,24] Patients with sepsis require emergency surgery. They constitute a progressively aging population using immunomodulatory agents and undergoing immunosuppressive therapies for cancer treatment. These patients are susceptible to secondary infections and potential complications that ultimately increase perioperative morbidity and mortality rates.^[25] A postmortem study revealed that most patients with sepsis had unresolved septic foci at the time of death. This indicates that the immune system of patients with sepsis is compromised and unable to eliminate the invading pathogens.^[26] Immunosuppressive properties of septic immune responses are increasingly becoming pertinent with advancements in critical care, because many deaths due to sepsis do not manifest acutely but instead occur following admission to an intensive care unit postoperatively or/and an extended hospitalization.^[23] During the different stages or for different types of perioperative sepsis, the primary objective of immunotherapy, especially for those in a preoperative immunosuppressed state, is regulating the host’s immune balance. Increasing evidence of reversing specific defects in late sepsis through immune stimulation strategies, including the administration of granulocyte-macrophage colony-stimulating factor (GM-CSF), suggests that the optimal immunotherapeutic approach may vary considerably depending on the stage of sepsis.^[27,28] However, anesthesiologists usually have limited time to systemically evaluate these patients before operation and adjust their immune microenvironment toward the most suitable state for operation. Due to severe tissue damage, drugs that are metabolized are not retained in the local area for a long time, and the treatment of patients with sepsis undergoing surgery is limited.

Therapeutic strategies primarily target two phases of sepsis: excessive inflammation and immune suppression. Unfortunately, none of them have shown substantial clinical benefits.^[23,29] The discrepancy between the current understanding of the pathophysiological processes and the lack of success in drug development for sepsis suggests that a single-target therapy is unlikely to be effective. Given the rapid progress in chimeric antigen receptor (CAR)-T cell therapy for cancer treatment,^[30] cell therapy is promising for the treatment of sepsis. Numerous pre-clinical and clinical studies have demonstrated the potential benefits of cell-based therapies in personalized treatment of sepsis.

Mesenchymal Stem Cell (MSC)-based Therapy Holds Promise for the Treatment of Sepsis

Since the early 1980s, cell-based therapeutic strategies have been developed and shown remarkable progress in recent years. The latest advances in stem-cell technologies for genetic and neurodegenerative diseases, and CAR-T therapy for cancer, have garnered global interest for their application in the treatment of sepsis.^[31,32,33]

Cell therapy involves the transfer of autologous or allogeneic cellular material to a patient for medical purposes.^[34] Critically ill patients often suffer multiple organ failures, including injuries to the lungs, kidneys, liver, or brain in the late stages. Once these patients undergo emergency surgery, the risk of severe infections and sepsis is high. Simultaneously, the negative effects of anesthesia and anesthetic agents exacerbate the impairment of systemic tissue perfusion and oxygenation, resulting in the collapse of cardiovascular function and respiratory dysfunction. This can lead to multiple organ dysfunction syndrome (MODS) and even death. As a potential therapeutic strategy, cell-based therapy is crucial in repairing local damage to vital organs and restoring homeostasis under sepsis state.^[8,23] Recent years have witnessed exponential advancements in pre-clinical and clinical trials focusing on cell-based therapies.^[35] As a research focus, several clinical trials (phases I and II) have demonstrated the safety of MSCs in acute respiratory distress syndrome (ARDS) and other diseases.^[36,37,38] Intraoperative stem cell therapies offer a prompt disease-modifying therapeutic strategy for the restoration of injured tissues and organs. Non-stem cell-based therapies, including natural killer (NK) cells, T cells, B cells, and macrophages, are ideal candidates for cellular engineering in the development of immunotherapy for sepsis.^[39]

MSCs

MSCs, also known as multipotent stromal cells or mesenchymal stromal cells, possess multi-directional differentiation potential. They can differentiate into osteocytes, adipocytes, and chondrocytes in vitro.^[40] MSCs are traditionally isolated from bone marrow but are found in numerous other sites including adipose, lung, heart, muscle, and fetal tissues.^[41] Given the low expression of cell-surface human leukocyte antigen class I and II molecules in MSCs, these cells can be used in both allogeneic and autologous therapies without immunosuppression.^[42] MSCs possess significant immunomodulatory properties, have low immunogenicity, and are easy to isolate and propagate.^[43] Therefore, these cells are considered immune privileged and have emerged as a promising source for the treatment of several diseases.

Immunomodulatory properties of MSCs in sepsis

Multiple experimental models have demonstrated the therapeutic potential of MSCs in sepsis.^[44,45,46] In several preclinical sepsis models, MSCs have exhibited anti-inflammatory effects, such as decreasing the production of pro-inflammatory cytokines (IL-6, tumor necrosis factor alpha [TNF-α], and interferon [IFN]-γ), increasing that of anti-inflammatory cytokines and molecules (IL-1 receptor antagonist, IL-10, cyclooxygenase-2, and prostaglandin E2 [PGE2]), and reducing the influx and accumulation of neutrophils.^[44,45] MSCs can modulate inflammatory responses and protect organ function during sepsis [Figure 4]. A potential mechanism by which MSCs can alter inflammatory responses in sepsis is through targeting TLR4 and inhibiting the activation of the nuclear factor kappa B (NF-κB) pathway. Another mechanism involves the reprogramming of host macrophages by MSCs through PGE2, resulting in increased production of anti-inflammatory cytokines including IL-10 by macrophages.^[46] When administered intravenously, MSC treatment can alter both local and systemic cytokine profiles during lipopolysaccharide (LPS)-induced endotoxemia. This can lead to reduced local inflammation and improved organ function.^[47] MSCs inhibit the mitochondrial apoptosis pathway, exert antioxidative effects, and modulate the expression of apoptosis-related proteins (caspase-3 and caspase-7).^[48] These properties represent another mode by which MSCs protect against immune and host cell damage induced by sepsis. MSCs also enhance macrophage-mediated phagocytosis and bacterial clearance by promoting CD206 expression through an IL-6-dependent mechanism.^[49] Numerous studies have confirmed the potential efficacy of MSCs in increasing survival in mouse models of sepsis induced by cecal ligation and puncture (CLP).^[50]

Figure 4.

Potential therapeutic efficacy of MSCs for sepsis. MSCs are sampled from the bone marrow, adipose tissue, umbilical cord of fetal tissue, muscles, and other organs. Their immunomodulatory capacities, owing to the secretion of several molecular factors, are used for the treatment of sepsis. MSCs control inflammation by reducing the levels of proinflammatory cytokines, increasing those of anti-inflammatory cytokines, and decreasing neutrophil and macrophage infiltration into damaged tissues. MSCs exert antimicrobial effects as they augment innate immune cell-mediated phagocytosis and bacterial clearance. MSCs can interact directly or indirectly through different cellular processes, such as cell differentiation and proliferation, angiogenesis, anti-apoptosis, and mitochondrial transfer during the progression to multi-organ dysfunction during sepsis. Given their various properties, MSCs can promote tissue repairing and organ function recovery, thus conferring a therapeutic benefit for sepsis. DC: Dendritic cell; MSCs: Mesenchymal stem cells; NK: Natural killer.

Anesthesia outlook on MSC application in sepsis

Septic patients often undergo an open surgical procedure to control infection source rapidly and adequately. However, it is not always effective. Surgical interventions may cause further release of infective molecules systemically. The promising clinical use of MSCs has been highlighted in many surgical procedures, including thoracic, cardiac, orthopedic, and abdominal surgery. Septic patients may show decreased potential morbidity following MSC-based treatments when surgical manipulations are urgent. MSCs can move and implant at a trauma site. They secrete anti-inflammatory mediators and wound-healing factors through the paracrine pathway to repair damaged tissues. These cells also aid in the pathogen clearance in infected tissues directly through phagocytosis and indirectly through the modulation of the function of patients’ phagocytes.^[51] Tracheal administration of MSCs may facilitate the repair of alveolar epithelial cell, thereby improving pulmonary ventilation/gas exchange dysfunction and ultimately alleviating hypoxia in septic patients during surgery. Conventionally, MSCs can directly be delivered into trauma or infected sites either systemically or by local injection using a suitable scaffold. In recent years, many bioengineering approaches have been developed to increase therapeutic efficacy, thus facilitating better control over the cell niches and regulation of their phenotype and function.

Several studies have demonstrated a significant improvement in cardiac function in patients who directly received intra-myocardial injections of MSCs. The treatment is safe over a long follow-up.^[52] MSCs play a role in mitigating the coagulation cascade, restoring endothelial barrier function, and promoting angiogenesis during the progression of multi-organ dysfunction during sepsis.^[53] Therefore, MSCs may represent a unique opportunity to reverse sepsis-induced circulatory failure, respiratory dysfunction, and coagulation dysfunction. Phases I and II clinical trials have been conducted to evaluate the safety and effectiveness of MSCs in the treatment of sepsis. Ten clinical trials to investigate MSCs for the treatment of sepsis have been registered on https://www.clinicaltrials.gov, among which three have been completed so far [Table 1]. Alp et al^[50] confirmed that no deaths resulted from allogenic adipose MSC infusions in 10 septic patients; five doses per patient were administered, suggesting a positive effect on sepsis in the early phase. McIntyre et al^[54] conducted an open-label phase I clinical trial to evaluate the safety of different doses of MSC therapy. Infusion of high-dose allogeneic bone marrow-derived MSCs into patients with septic shock was found to be safe. In a phase I clinical trial, Perlee et al^[55] determined the effect of MSCs in response to intravenous LPS stimulation in healthy subjects. They confirmed that infusion of allogenic adipose MSCs exerted anti-inflammatory and pro-coagulant effects. Only one phase I clinical trial in China by He et al^[56] is registered on https://www.chictr.org.cn. They reported the safety and efficacy of transplantation of MSCs from the human umbilical cord in patients with sepsis. The mortality rate was 20% (3/15) in septic patients who received a single-dose MSC treatment, lower compared with the expected mortality rate due to sepsis according to the global statistics. However, the small sample sizes of these studies undermine their credibility. Larger sample sized phase III trials are needed to define the adverse events of MSC-related therapies.

Table 1.

Cell therapy-based clinical trials for sepsis.

Cell types	NCT numbers	Titles	Conditions	Dose and route	Status	Phases	Countries	Results
MSCs	NCT05283317	Effect of mesenchymal stromal cells on sepsis and septic shock	Sepsis/septic shock	1 × 106/kg	Completed	Phase I/phase II	Turkey	Alp et al[50]
	NCT02883803	Treatment of severe infections with mesenchymal stem cells (CHOCMSC)	Septic shock	1 × 106/kg	Unknown	Not applicable	France	–
	NCT01849237	Russian clinical trial of mesenchymal cells in patients with septic shock and severe neutropenia	Septic shock	1–2 millions·kg–1·day–1	Unknown	Phase I/phase II	Russia	–
	NCT04961658	Advanced Mesenchymal Enhanced Cell THerapY for SepTic Patients (AMETHYST)	Septic shock	15 or 60 or 150 million cells	Recruiting	Phase I	Canada	–
	NCT02421484	Cellular immunotherapy for septic shock: A phase I trial (CISS)	Septic shock	0.3 or 1.0 million or 3.0 million cells/kg intravenously	Completed	Phase I	Canada	McIntyre et al[54]
	NCT02328612	Randomized, parallel group, placebo control, unicentric, interventional study to assess the effect of expanded human allogeneic adipose-derived mesenchymal adult stem cells on the human response to lipopolysaccharide in human volunteers (CELLULA)	Sepsis	0.25 or 1 million or 4 million cells/kg intravenously	Completed	Phase I	Netherlands	Perlee et al[55]
	NCT02789995	Dysfunctions of human muscle stem cells in sepsis (DISCUSS)	Sepsis	Not known	Completed	Not applicable	France	Duceau et al[57]
	NCT05969275	Umbilical mesenchymal stromal cells as cellular immunotherapy for septic shock (UC-CISSII)	Sepsis/septic shock	300 million cells intravenously	Not yet recruiting	Phase II	Not applicable	–
	NCT03369275	Cellular immunotherapy for septic shock (CISS2)	Sepsis/septic shock	300 million cells intravenously	Unknown	Phase II	Not applicable	–
	NCT04445220	A study of cell therapy in COVID-19 subjects with acute kidney injury who are receiving renal replacement therapy	Sepsis	250 or 750 million cells	Unknown	Phase I/phase II	United States	–
Immune cell	NCT05442710	Recovery from acute immune failure in septic shock by immune cell extracorporeal therapy (ReActiF-ICE)	Sepsis	Not known	Recruiting	Phase II	Germany	–
Granulocyte	NCT00818597	Extracorporeal immune support system (EISS) for the treatment of septic patients (EISS-1)	Sepsis/septic shock	Not known	Completed	Phase I/phase II	Germany	Altrichte et al[58]
Monocyte/apoptotic cells	NCT03925857	Prevention of sepsis-related organ dysfunction with apoptotic cells (Allocetra-OTS, Enlivex Therapeutics Ltd, Tel Aviv, Israel) (P-SOFA-1)	Sepsis	One dose Allocetra-OTS 140 × 106/kg	Completed	Phase I	Israel	van Heerden et al[59]
	NCT04612413	A phase 2 study evaluating efficacy, safety and tolerability of different doses and regimens of Allocetra-OTS for the treatment of organ failure in adult sepsis patients	Sepsis	Single dose of Allocetra-OTS, 5 × 109 or 10 × 109 cells intravenously	Recruiting	Phase II	Belgium	–
T cells	NCT02136797	Trial of third party donor derived CMVpp65 specific T-cells for the treatment of CMV infection or persistent CMV viremia after allogeneic hematopoietic stem cell transplantation	Viremia	1 × 106 cells·kg–1·dose–1·week–1 intravenously	Recruiting	Phase II	United States	–
	NCT03798301	Treatment of cytomegalovirus (CMV) infections with viral-specific T cells	Viremia	25,000 cells/kg intravenously	Recruiting	Phase I	United States	–
	NCT04605484	Study of Posoleucel (formerly known as ALVR105; Viralym-M) in kidney transplant patients with BK viremia	Viremia	Not known	Completed	Phase II	United States	–
	NCT01646645	Primary transplant donor derived CMVpp65 specific T-cells for the treatment of cmv infection or persistent CMV viremia after allogeneic hematopoietic stem cell transplantation	Viremia	1 × 106 cells·kg–1·dose–1·week–1 intravenously	Completed	Phase II	United States	–
	NCT04018261	Virus-specific activated T lymphocytes from a donor in hematopoietic progenitor transplanted patients	Viremia	Not known	Active, not yet recruiting	Phase I/phase II	Spain	–
Lymphocyte	NCT06011486	Expansion of virus-specific lymphocytes for cell therapy	Viremia	Not known	Recruiting	Phase I	Brazil	–
NK cell	NCT04320303	CMV infection and immune intervention after transplantation	Viremia	Not known	Unknown	Phase II	China	–
DCs	NCT05786937	Therapeutic vaccine based on aDC1 dendritic cells for the control of viremia after ATI in HIV infected individuals	Viremia	Not known	Not yet recruiting	Phase I	Brazil	–

DC: Dendritics cell; MSC: Mysenchymal stem cell; NCT: National clinical trial; NK: Natural killer; –: Not reported.

Several unknown parameters exist in the procedure of optimal delivery of MSCs, including the type of MSCs that needs to be infused, cell storage method (cell viability and if they should be fresh or frozen), dosing regimen (including dose, interval, and number of cycles), and route of administration. During surgery, the status of septic patients changes continuously. The optimal treatment strategy for the patients at the appropriate time needs to be determined. Addressing the above challenges is expected to greatly benefit the management of patients with sepsis.

Cell-based Immunotherapy for Sepsis

In clinical practice, patients experiencing sepsis-induced immunosuppression develop profound metabolic anomalies in their immune cells, prompting a deeper exploration of potential therapeutic interventions.^[8,23] Given that a significant proportion of septic patients requiring surgical intervention are older adults grappling with additional comorbidities and complications, including concurrent malignant tumors requiring immunosuppressive therapy, the clinical pathophysiology and perioperative management are intricate.^[60,61,62] Consequently, individualized therapeutic approaches and reinforcement of immune function are paramount for this unique population. Direct monitoring of immune cells is an exciting avenue for further investigation. Previous research centered on CAR-immune cell therapy for cancer has demonstrated the potential of reprogramming immune cells as a promising therapeutic strategy for various diseases.^[30,32] Drawing inspiration from this, several studies have attempted to monitor immune cells in sepsis. In this context, our primary focus centers on the role of macrophages in the adaptive immune system. Other immune cell types in the context of sepsis have been briefly discussed.

Macrophages

Macrophages, the primary natural immune cells, and key antigen-presenting cells, substantially affect both innate and adaptive immunity.^[63] Occupying the role of resident sentinels within the body, macrophages are crucial in the body’s defense against pathogens and immunopathogenesis during the initial stages of infections and sepsis. Circulating macrophages respond rapidly toward bacterial stimulation following surgical trauma. During sepsis, they participate in the process of pathogen phagocytosis, cytokine secretion, and phenotype reprogramming, and clear tissues of worn-out cells, damaged organelles, and debris upon tissue damage.^[11] In response to diverse environmental cues (such as microbial products and damaged cells) and the unique microenvironment of a specific tissue type, macrophages can transform into different phenotypes, each demonstrating distinct functions in sepsis.

Previously, macrophages were typically classified into two main types, namely M1- and M2-like macrophages.^[63] The pro-inflammatory M1 phenotype, or classically activated macrophages, is found when monocyte-derived macrophages are stimulated with IFN-γ or bacterial LPS. These cells produce massive amounts of inflammation-related mediators that recruit other inflammatory cells, like neutrophils, to reinforce the host’s immune defense against invading pathogens.^[64] Conversely, anti-inflammatory M2 macrophages, also known as alternatively activated macrophages, are generated following stimulation with IL-4/13. They are recognized for their regulatory role in tissue remodeling, repair, and angiogenesis. M2 macrophages modulate inflammatory responses, clear apoptotic cells, and facilitate wound healing. The traditional classification of macrophages (M1 and M2) is unclear on the cellular compartmentalization, which limits its application to different immune or tissue microenvironments in sepsis.

With the development of lineage tracing, single-cell sequencing, and cell fate mapping technologies, macrophages were found to comprise distinct subpopulations with diverse origins, functional characteristics, and transcriptional regulatory mechanisms that are tissue or disease-specific.^[65] The transcriptional profile of human macrophages exposed to multiple signals has revealed a spectrum of activation states of macrophages that extend the classic concept of macrophage M1/M2 polarization.^[65] The generation of transcriptomic datasets of human macrophages has broadened the understanding of integration of signals from the local microenvironment by macrophages.^[66] Manufactured CAR macrophages can redirect phagocytic function and trigger an adaptive immune response. They have entered the clinical evaluation stage and show promise in the treatment of diseases.^[67,68] In several diseases, such as tissue injury, inflammatory disease, degenerative disease, organ fibrosis, and tumors, ex vivo-generated macrophages, as cell-based therapies, have outperformed stem cells.^[67] Research on CAR macrophages is at the pre-clinical stage with one ongoing phase I trial based on autologous CAR macrophages targeting human epidermal growth factor 2 (HER2)-overexpressing solid tumors.^[67] Remarkable progress has been made in the utilization of macrophage-based immunotherapy for protection against multiple organ failure in sepsis. Most septic patients suffer from cardiac dysfunction which significantly contributes to hemodynamic instability and hypoperfusion during transfer for operation. Using a mouse model of CLP-induced sepsis, a subpopulation of cardiac-resident triggering receptor expressed on myeloid cells (TREM) 2^high macrophages (termed Mac1) was identified. These protected the septic heart by actively scavenging dysfunctional mitochondria ejected from cardiomyocytes.^[69] Intriguingly, the transfer of TREM2^high macrophages to the pericardial cavity immediately showed recovery cardiomyocyte function and improved effective perfusion of microcirculation, convincingly suggesting that cell-based immunotherapy is effective for sepsis and subsequent cardiac dysfunction. The recovery of cardiac function results in suitable surgical conditions and physiological status for septic patients. Liver injury in sepsis is reduced through the regulation of Kupffer cells’ activity, further improving bacterial clearance and outcome in septic cases.^[70] Park et al^[71] demonstrated that kidney-resident macrophages express V-domain Ig suppressor of T cell activation (VISTA), an injury-repairing factor. It can accelerate the process of post-ischemic renal repair by scavenging apoptotic cells and controlling T cell infiltration. Bang et al^[72] confirmed that artesunate and neuroprotectin D1 initiate the activation of GPR37 in macrophages that can confer protection against infection-induced sepsis. Suzuki et al^[73] used a mouse model of GM-CSF receptor-β-deficiency (Csf2rb^–/–). They found that GM-CSF is required for phenotype conversion of Csf2rb-gene-corrected macrophages in the lung environment. Pulmonary macrophage transplantation of Csf2rb-gene-corrected macrophages reversed the abnormal surfactant accumulation for at least one year. These experimental findings imply that the tissue microenvironment is crucial in determining the macrophage phenotype and function.

Manifestations of sepsis depend on a complex interplay between the host, pathogen, and environmental factors. In sepsis, macrophages participate in a complex interaction with the host immune system owing to their unique features, including outstanding propensity of migration and infiltration, phagocytic ability, and inflammatory modulation. Macrophages show different functions and phenotypes—harmful or beneficial—depending on the immune microenvironment of the specific organ where they reside and at different stages of sepsis. Targeting specific macrophage sub-populations to resolve injured organs and promote organ function recovery during sepsis bears more perioperative therapeutic value.^[11,65] Although only in the preclinical stage, macrophages, as immune cells that are rapidly mobilized perioperatively as the first line of defense, show advantages distinct from MSCs. While numerous questions must be addressed, the accumulation of additional pre-clinical data and clinical evidence is anticipated to extend their applications to novel therapeutic frontiers.

Neutrophils

Neutrophils, originating from hematopoietic cells of the bone marrow, constitute a vital population of leukocytes crucial to innate immunity.^[74] Serving as the foremost line of defense against invading pathogens, neutrophils primarily function through processes like digestion, phagocytosis, and bactericidal activity.^[74] Mounting evidence suggests that neutrophils have a dual-edged role in sepsis. While they represent the initial wave of leukocytes entering the peripheral blood, neutrophils can exhibit aberrant functions during the surgical stage of sepsis, including delayed apoptosis, anomalous migratory chemotaxis, formation of extracellular traps, and release of human neutrophil defensin. Consequently, these anomalies contribute to the dysregulation of host immune responses, coagulopathy, endothelial cell pyroptosis, and a decline in organ perfusion.^[75,76] Monitoring neutrophil activity is a potential method for detecting sepsis.^[77] No clinical trials based on neutrophils are registered on https://www.clinicaltrials.gov. Further in-depth research is essential to unravel the intricate mechanisms underlying the role of neutrophils in sepsis.

NK cells

NK cells, which are pivotal antiviral lymphocytes of the innate immune system, contribute substantially to the overall host immunity.^[78] Their significance as mediators in sepsis was first recognized in the latter half of the 20th century. Heightened NK cell activity can potentially ameliorate viral infections, including severe infection-induced sepsis.^[79] However, as evidenced in animal studies, this cell type exerts diverse effects during sepsis. On the one hand, they bolster host resistance to infection by secreting key factors such as IFN-γ, GM-CSF, and TNF, along with other anti-inflammatory mediators including antimicrobial peptides and α-defensins. On the other hand, NK cells have been implicated in deleterious roles in sepsis, such as exacerbating inflammatory responses and tissue injury, ultimately leading to organ dysfunction.^[80] Giamarellos-Bourboulis et al^[81] found that elevated NK cell counts were associated with increased survival rates in over 20% of septic patients, which was attributed to elevated levels of soluble triggering receptor expressed on myeloid cells-1 (sTREM-1) in their sera. Other studies yielded contrasting results, which suggest that depleting NK cells in mice could enhance their tolerance to sepsis.^[82,83] No clinical trials centered on NK cell therapy in sepsis have been conducted. In a recent study, Lu et al^[84] employed a mouse model with SARS-CoV-2 infection to generate CAR-NK cells using the mutant extracellular domain of angiotensin converting enzyme 2 along with human soluble IL-15. These modified cells extended their lifespan, hinting at their potential applicability in the treatment of infectious diseases.

Taken together, despite the intricate role of NK cells in sepsis, their current clinical application remains limited. Nonetheless, their robust capacity to combat virus-infected cells holds promise for the mitigation of perioperative sepsis. Although an adequate study in this area is lacking, discrepancies open up avenues for further investigation. NK cells bear the potential to serve as a fundamental immunotherapeutic strategy, encompassing in vitro expansion and various types of CAR approaches for patients with sepsis. The emerging field of NK cell and CAR-NK cell-based immunotherapy for bacterial infections warrants increased research consideration.

DCs

DCs are specialized antigen-presenting cells with a crucial bridging role in the initiation and regulation of innate and adaptive immune responses. They secrete cytokines such as IL-12 and IFN-α and interact with various immune cells. During sepsis, the number of DCs decreases and their function reduces remarkably. This cellular loss is more prominent in patients with sepsis showing worse clinical outcomes.^[85] Enhancing DC function can improve outcomes in animals with sepsis by mitigating immune suppression.^[86] Regulating DC numbers and functions is a potential target in the treatment of sepsis. Clinical trials based on DCs are currently recruiting patients, and the results are unknown. More evidence of targeting DCs, both pre-clinical and clinical, can shed light on the treatment of perioperative sepsis.

T cells and B cells

T cells, an important part of adaptive immunity, drive and control immune responses. T helper (Th)1 and Th2 cells are involved in an interplay and the balance between them plays a role in infection resolution. During immunosuppression in sepsis, CD4⁺ T cells, CD8⁺ T cells, Th17 cells, and γδT cells decrease, while regulatory T (Treg) cells increase.^[85] T cell exhaustion is a significant feature during the prolonged stage of sepsis which is strongly associated with immunosuppression.^[12] Immune metabolism is more likely to determine the fate and immune function of T cells.

Various clinical studies have reported suppressed T cell function, increased T cell apoptosis, and diminished secretion of Th1- and Th2-associated cytokines in the late stage of sepsis.^[87] Along with the up-regulation upon onset of sepsis, the persistently dynamic increase in Th1/Th2 is associated with intensive care unit-acquired infection and death. These results can help develop a precisely stratified adaptive immune-targeted therapy.^[88] There is an imbalance of Th17/Treg cells in patients with sepsis that can be attenuated by high-volume hemofiltration. Pre-clinical studies based on T cells indicate a prospective therapy for patients with sepsis. For example, by using Treg cells from healthy mice and transferring them to mice with acute kidney injury, Lu et al^[89] demonstrated the effect of these cells in restoring the function and anatomical structure of the injured kidney. Given that Treg and Th17 cells are distinct CD4⁺ T cell subsets from Th1 and Th2 cells, Guo et al’s^[90] study showed that Th17, Treg, Th17/Treg levels and production of cytokines were significantly higher in patients with sepsis; high-volume hemofiltration could attenuate the Th17/Treg imbalance and improve organ dysfunction. In a sepsis-associated encephalopathy (SAE) model induced by polymicrobial, Saito et al^[91] found an increased number of infiltrated Treg cells and Th2 cells in the brain. These cells could attenuate SAE and SAE-induced mental deterioration by modulating neuroinflammation. These results suggest that the T cell-based therapeutic strategy is a potential direction for the treatment of sepsis.

B cells exhibit multiple functions. Along with changes in different B cell sub-populations, increased apoptosis of memory B cells, persistent decrease in primitive B cells, and an increase in B cell depletion occur during the development of sepsis.^[92] In patients with septic shock during intensive care unit stay, expressions of CD23⁺, CD95⁺, or CD80⁺ in B cells were found to be significantly associated with higher mortality.^[93] Other studies have reported that the numbers of circulating B cells and serum immunoglobulin M levels are significantly associated with poor prognosis of patients with sepsis.^[94]

However, emerging evidence suggests that B cells may play a protective role in pathogen clearance and an immune regulatory role in sepsis. Previous studies have primarily delved into the underlying mechanisms of B cell function in the context of sepsis-induced systemic inflammatory response. B cells enhance pathogen clearance and cytokine production through the type I IFN signaling pathway when eliciting initial innate immune responses, signifying a protective role in sepsis.^[95] In a pre-clinical model employing septic mice, Rauch et al^[96] identified a subset of effector B cells, termed innate response activator B cells. Remarkably, these B cells, derived from B1a B cells, showed a protective effect against septic shock by reducing bacterial clearance, mitigating cytokine storms, and minimizing organ injury. When combined with evidence indicating that reduced numbers and impaired functionality of B cells are common in sepsis and are linked to poor outcomes, restoring B cell numbers and function may bear the potential for improving the prognosis of patients with sepsis. Current B-cell-based immunotherapy primarily focuses on mitigating B-lymphocyte loss and supplementing B cells and immunoglobulins G, A, M (IgGAM) in vitro.^[97,98] More clinical trials centered on T- and B-cell-based therapy in sepsis are expected in the future.

Clinical Practice and Challenges to Cell-based Immunotherapy in Perioperative Sepsis

In clinical settings, patients with sepsis share similar pathophysiological processes and succumb to multi-organ failure, due to the intricate interplay of overwhelming inflammation and immunosuppression. However, the clinical course of septic patients undergoing surgery is different from that of patients whose disease progresses in the intensive care unit or internal ward. Septic patients predominantly show infection caused by Gram-negative bacteria and surgical removal of the infection is the most successful method. From the perspective of anesthesia or perioperative period, sepsis is heterogeneous. Susceptible organ damage involved before operation and during operation for sepsis is different. Therefore, comprehensive preoperative evaluation, close intraoperative monitoring, timely intervention, and focused postoperative recovery and management are urgent in anesthetic care for these high-risk patients.

Notable in clinical trials are MSCs, advancing into phase II research owing to their immune regulation and tissue repair capabilities. These cells have shown promise in addressing the imbalance of ventilation and blood flow ratios and reducing intrapulmonary shunt in patients with sepsis during surgery.^[55,56] Tracheal administration of MSCs may facilitate the repair of alveolar epithelial cell, thereby improving pulmonary ventilation/gas exchange dysfunction and ultimately alleviating hypoxia in septic patients.

The evolution of single-cell sequencing technology has shed light on the important role of immune cells, especially various macrophage subpopulations, in maintaining homeostasis and tissue repair. These subpopulations, characterized by distinct gene expression profiles, can adapt their phenotypes in response to environmental signals. In recent years, disease-specific mature macrophage subpopulations have garnered greater attention compared to MSCs and other innate immune cells. They are essential for the clearance of cellular debris, phagocytosis, and tissue repair, ultimately maintaining homeostasis.^[11] The profound and dysregulated activation of macrophage subgroups during sepsis may exert a direct effect on its outcome. TREM2^high macrophages scavenge dysfunctional mitochondria ejected from cardiomyocytes to protect septic heart.^[69] In cases of heart failure identified preoperatively where cardiotonic agents and vasodilators are ineffective, the use of TREM2^high macrophages via pericardial injection may represent a new approach to promoting the recovery of the heart pump function, thus facilitating smoother surgical procedures for sepsis patients. It is conceivable that the protective effects seen in the heart also extend to other tissues where specific macrophages subpopulations ubiquitously distributed are also shown to restore tissue homeostasis. Disruption of mucosal barrier permeability occurs more frequently in these patients with compromised physical status and host immune disturbance. Gut-resident macrophages (GRMs) are crucial for the regulation of the immunological microenvironment in the gastrointestinal system and the promotion of barrier function. The induction of monocyte differentiation into GRMs by activating transcription factor 4 mitigates inflammation in a murine sepsis model, which can be potentially adapted to the management of sepsis-related gastrointestinal dysfunction.^[99] Postoperative care, conventionally dependent on antibiotics and nutrition, could benefit from targeted macrophage administration, potentially speeding up recovery and improving long-term prognosis. This approach may promote the recovery of intestinal function, reduce hospital stays, and improve long-term prognosis.

Cell-based therapies for sepsis management are important given the unique features of surgical patients. Mechanisms underlying these therapeutic approaches include targeted recruitment and differentiation for functional reprogramming of the activated macrophages through activation or inhibition of specific receptors by scRNA-seq and genetic engineering. Despite our lack of understanding regarding the mechanism, it is appealing to modify the cells into diverse phenotypes based on particular pathological processes in septic patients. However, the challenges are prominent. First, the realization of precise modification of immune cells requires cutting-edge techniques. Second, adequate clinical evidence to demonstrate the safety and efficacy of cell therapies is lacking and warrants further evidence. Administration of immune cells may elicit immunological rejection, which can subsequently trigger severe cascade responses in these patients. Different delivery approaches affect the efficiency of cell-based therapies. However, the influencing factors are elusive and need further investigation. High costs of cell-based therapies should be considered, as they pose a heavy financial burden on the patients.

The advancements in gene editing and haploidentical bone marrow transplantation are also instrumental to creating modified immune cells and overcoming donor limitations, respectively.^[100] This progress suggests that obtaining macrophages from allogeneic sources could be a future trend. Further research focusing on understanding the mechanisms of cell therapy in sepsis, along with verifying its efficacy and safety, is crucial for its clinical implementation. This could mark a new era in the treatment of sepsis patients undergoing surgery.

Conclusions

Sepsis presents a significant challenge for anesthesiologists. The implementation of bundled therapies, including early fluid resuscitation, prompt administration of antibiotics, hemodynamic stabilization, and organ-supportive interventions, has substantially enhanced patient survival. However, a definitive solution for efficiently addressing multi-organ failure induced by sepsis remains lacking. Considering the available evidence, we posit that novel cell-based immunotherapy holds promise as a new avenue for the treatment of sepsis. MSC-based therapies, particularly those derived from adipose tissue, are gaining traction as a potential treatment for sepsis, with adipose-derived MSCs already advancing to the phase II clinical trial stage (No. NCT05283317). Future studies should delve deeper into aspects such as optimal dosage, administration route, and storage methods. Macrophages, while still in the pre-clinical stages for the treatment of sepsis, have demonstrated significant potential. Specifically, TREM2^high macrophages exhibit proficiency in clearing debris, facilitating tissue repair, and sustaining perfusion, ultimately modulating the outcome of cases of surgical sepsis. Given their robust phagocytic and bacterial defense abilities, macrophages hold promise as a therapeutic strategy for the treatment of sepsis. Other immune cells, including neutrophils, NK cells, DC cells, T cells, and B cells, also exhibit potential in the treatment of perioperative sepsis. The evolving landscape of cell therapy in treating perioperative sepsis signifies a breakthrough in medical science. Continued research into these mechanisms as well as their efficacy and safety are stringent for clinical implementation, heralding a new chapter in sepsis treatment for surgical patients.

Acknowledgment

We thank Chuanbin Mao from School of Materials Science & Engineering, Zhejiang University for his help with linguistic refinement and edition of this article.

Funding

This work was supported by a grant from the National Natural Science Foundation of China (No. 82230074).

Conflicts of interest

None.