Peripheral Injection of hUC-MSCs in the Treatment of Acute Liver Failure: A Pre-Clinical Cohort Study in Rhesus Monkeys

Abstract

Background: Using a toxin-induced lethal acute liver failure (ALF) monkey model, we have recently shown that early peripheral infusion of human umbilical cord mesenchymal stem cells (hUC-MSCs) can alleviate liver damage and improve animal survival by suppressing the activation of circulating monocytes and the subsequent cytokine storm. Here, we explored whether the administration of hUC-MSCs could still improve ALF when the cytokine storm is fully developed.

Method: We treated ALF monkeys with peripheral delivery of hUC-MSCs at 48 hr after toxin challenge. Liver indices, histology, imaging, and animal survival were recorded and analyzed.

Results: In our cohort study, we conducted and demonstrated that the infusion of hUC-MSCs significantly improved liver histology, effectively controlled inflammatory cytokine storms, and increased survival rates. Additionally, the administration of a higher dose of hUC-MSCs (2 × 10⁷/monkey) yielded superior outcomes compared to a lower dose (1 × 10⁷/monkey).

Conclusion: Treatment of hUC-MSCs can significantly improve the pathological and survival outcomes of ALF even when the cytokine storm has been fully developed, indicating a promising clinical solution for ALF.

Figure 1

hUC-MSCs improve survival in monkeys with ALF. (a) Schematic representation of the experimental grouping. (b) Schematic representation of the experimental design. (c) Survival curves for the monkeys submitted to different treatments (Kaplan–Meier method with log-rank test) P < 0.001.

Figure 2

Peripheral delivery of hUC-MSCs ameliorates liver histology and hepatic indices in 20 μg/kg of α-amanitin group. (a) Biochemical assay of hepatic indices: alanine aminotransferase (ALT), glutamic-oxaloacetic transaminase (AST), total bilirubin (TBIL), prothrombin time (PT), activated partial thromboplastin time (APTT), and blood ammonia (BA). Error bars, SEM.  ^∗P < 0.05,  ^∗∗P < 0.01, and  ^∗∗∗P < 0.001, compared with the data at 48 hr. (b) HE staining and Oil red O staining of liver biopsy at indicated times.

Figure 3

Peripheral delivery of hUC-MSCs ameliorates liver histology and hepatic indices in 40 μg/kg of α-amanitin group. (a) Biochemical assay of hepatic indices: alanine aminotransferase (ALT), glutamic-oxaloacetic transaminase (AST), total bilirubin (TBIL), prothrombin time (PT), activated partial thromboplastin time (APTT), and blood ammonia (BA). Error bars, SEM.  ^∗P < 0.05,  ^∗∗P < 0.01, and  ^∗∗∗P < 0.001, compared with the data at 48 hr. (b) Gross and histopathological changes in a severe ALF liver. (c) Oil red O staining of liver biopsy at indicated times. (d) HE staining of liver biopsy at indicated times.

Figure 4

hUC-MSCs inhibit the activation of circulating monocytes. (a and b) Flow cytometry analysis of activated monocytes (CD14+ CD16+ CCR2+) in subsets of peripheral blood monocytes. Error bars, SEM, Student's t-test,  ^∗∗P < 0.01, and  ^∗∗∗P < 0.001.

Figure 5

hUC-MSCs suppress systemic inflammation. (a and b) Show the serum levels of cytokines including TNF-α (tumor necrosis factor), IL-6, IL-1β, IL-10, IL-4, and IL-1RA (IL-1 receptor antagonist) in response to 20 μg/kg and 40 μg/kg of α-amanitin challenge, respectively. Error bars, SEM.  ^∗P < 0.05,  ^∗∗P < 0.01,  ^∗∗∗P < 0.001, and  ^∗∗∗∗P < 0.0001, compared with the data at −48 hr.

Figure 6

hUC-MSC infusion promotes liver regeneration. (a) Ki67 immunohistochemical staining of liver specimens at indicated times after infusion of hUC-MSC. (b) Quantitation of Ki67+ positive hepatocytes at indicated times following different treatments. The numbers of positive cells in 10 consecutive high-power fields were counted. Error bars, SEM, Student's t-test,  ^∗∗P < 0.01,  ^∗∗∗P < 0.001, and  ^∗∗∗∗P < 0.0001.