Clinical-grade mesenchymal stem cells derived from umbilical cord improve septic shock in pigs

Abstract

Background: Septic shock is the leading cause of death in intensive care units. The pathophysiological complexity of this syndrome contributes to an absence of specific treatment. Several preclinical studies in murine models of septic shock have shown improvements to organ injury and survival after administration of mesenchymal stem cells (MSCs). To better mimic a clinical approach in humans, we investigated the impact of randomized controlled double-blind administration of clinical-grade umbilical cord-derived MSCs to a relevant pig model of septic shock.

Methods: Septic shock was induced by fecal peritonitis in 12 male domestic pigs. Animals were resuscitated by an experienced intensivist including fluid administration and vasopressors. Four hours after the induction of peritonitis, pigs were randomized to receive intravenous injection of thawed umbilical cord-derived MSCs (UCMSC) (1 × 10⁶ UCMSCs/kg diluted in 75 mL hydroxyethyl starch (HES), (n = 6) or placebo (HES alone, n = 6). Researchers were double-blinded to the treatment administered. Hemodynamic parameters were continuously recorded. Gas exchange, acid-base status, organ function, and plasma cytokine concentrations were assessed at regular intervals until 24 h after the onset of peritonitis when animals were sacrificed under anesthesia.

Results: Peritonitis induced profound hypotension, hyperlactatemia, and multiple organ failure. These disorders were significantly attenuated when animals were treated with UCMSCs. In particular, cardiovascular failure was attenuated, as attested by a better mean arterial pressure and reduced lactatemia, despite lower norepinephrine requirements. As such, UCMSCs improved survival in this very severe model (60% survival vs. 0% at 24 h).

Conclusion: UCMSCs administration is beneficial in this pig model of polymicrobial septic shock.

Keywords: Clinical-grade; Mesenchymal stem cells; Septic shock; Umbilical cord.

Fig. 1

Characterization of UCMSCs. Mesodermic differentiation of UCMSCs. Clinical-grade UCMSCs differentiated into adipocytes (a) and osteocytes (b). Representative images are shown at × 10 magnification. Immunophenotypic analysis of UCMSCs by flow cytometry (c). UCMSCs presented the typical immunophenotype of MSCs. Negative cocktail includes CD34, CD45, CD11b, CD19, and HLA-DR markers. Results are shown as percentages of positive cells and are expressed as mean ± SEM. (n = 3)

Fig. 2

UCMSCs protect from sepsis-induced hypotension and tissue hypoxia. Evolution of (a) mean arterial pressure (MAP), (b) norepinephrine requirements, (c) SvO2, (d) arterial lactate concentration, (e) cardiac index (CI), and (f) heart rate over the 24-h study period. MAP and SvO₂ were higher in UCMSC-treated animals than controls; whereas, norepinephrine dose and lactate levels were lower. (n = 6 per group).^*p < 0.05; ^**p < 0.01;^***p < 0.001; ^****p < 0.0001 versus control group

Fig. 3

UCMSCs attenuate sepsis-induced organ failure. Lung function was monitored by assessing the PaO₂/FIO₂ ratio (a) and based on histology (d). Renal function was assessed by measuring plasma creatinine concentration (b), fluid balance (c), and by histological observation (e). For histological scoring, a score of 0 corresponds to an absence of anomaly, 1 reflects discrete anomalies, 2 corresponds to moderate anomalies, and 3 indicates severe anomalies. Histological images (hematoxylin-eosin staining, × 40 magnification) are representative of the respective conditions. ^*p < 0.05; ^**p < 0.01;^***p < 0.001; ^****p < 0.0001 versus control group

Fig. 4

UCMSCs have no antibacterial effect. Quantitative blood cultures at the time of death (a). The graph represents the medians and IQR of bacterial CFU (n = 6 per group). UCMSCs were co-cultured at different concentrations with 10⁴ to 10⁵ CFU of three species of bacteria commonly found in intestinal microbiota: Pseudomonas aeruginosa (b), Bacteroides fragilis (c), and Staphylococcus aureus (d) (n = 3 per group). CFU counts were determined by serial dilution and agar plating after 3 h of co-culture. Differences between groups were not statistically significant

Fig. 5

UCMSCs have no effect on systemic cytokine concentrations. Plasma concentrations of TNFα (a) and IL-6 (b) were similar between groups (n = 6 per group)

Fig. 6

UCMSCs improve survival. Survival percentages of untreated and treated pigs after induction of peritonitis are presented as a Kaplan-Meier survival curve (Log-Rank test, p = 0.08)