Immunomodulatory Plasticity Of Mesenchymal Stem Cells And Its Transformation In Rheumatoid Arthritis

BackgroundMesenchymal stem cells?MSCs?are a group of multi-potent adult stem cells that exist in many tissues and are capable of differentiating into several different cell types.Beside their active effect in tissue regeneration and repair,MSCs also possess immunoregulatory properties that can modulate immune as well as inflammatory responses.Exogenously administered MSCs migrate to damaged tissue sites,where they participate in tissue repair.Depending on their type and intensity,inflammatory stimuli confer on MSCs the ability to suppress the immune response in some cases or to enhance it in others.By regulating proliferation,differentiation,and functional status of immune cells,MSCs can regulate the secretion of inflammatory cytokines,showing great potential in the fields of regenerative medicine and immunotherapy in inflammation-related diseases.Although there have been many studies on the immunomodulatory capacity of MSCs,the regulation of the plasticity of MSCs in immunomodulation remains unclear,which hinders the development of new strategies to improve its clinical efficacy and the application of MSCs in immune-related diseases.Sepsis is a systemic excessive inflammatory response syndrome caused by infectious factors,often causing multiple organ dysfunctions.It is the leading cause of death in ICU patients.The mortality rate is as high as 50%⁷0%.At present,there is no effective treatment except hormone anti-inflammatory,antibiotic sterilization and symptomatic support therapy.At present,the comprehensive strategy of"bacteria,toxicity,inflammation and tissue repair"is considered to be the optimal treatment for sepsis.In addition to tissue regeneration and repair,MSCs also have strong anti-inflammatory and anti-bacterial capabilities,and animal experiments have confirmed that MSCs could significantly reduce the mortality of sepsis mouse model.Therfore,it is considered to be a new hope for the treatment of sepsis.However,the current clinical translation studies on MSCs for the treatment of sepsis have not been reported.Rheumatoid arthritis?RA?is a systemic autoimmune disease characterized by chronic aggressive synovitis.Hormones,anti-rheumatic drugs or biological agents are commonly used to treat RA in clinic.However,more than 50%of RA patients still fail to achieve clinical remission after treatment.In addition,although the above treatment measures can improve the clinical symptoms of arthritis in a short time,they still can not achieve the goal of long-term clinical remission or even cure.The long-term prognosis of RA patients is poor,and the disease often recurs.Therefore,exploring an effective strategy to cure RA is still a difficult problem in this field.In view of the strong anti-inflammatory and immune regulation capabilities of MSCs in addition to tissue regeneration and repair,the treatment of RA by MSCs can fundamentally regulate the imbalanced immune system and cure the disease.Aim1.To investigate the molecular mechanism of immunoregulatory plasticity?PD-L1expression?in MSCs stimulated by exogenous TNF-?combined with IFN-?.2.Pioneered a single-center,open-label,dose-escalation phase one clinical trial of a single dose of intravenous MSCs in patients with severe sepsis,systematically test the safety and feasibility of single dose allogeneic umbilical cord derived MSCs in pat ients with severe sepsis.3.To explore the mechanism or factors influencing the clinical efficacy of mesenchymal stem cell transplantation?MSCT?in the treatment of rheumatoid arthritis?RA?,and to provide reliable serum biomarkers for the patient inclusion and prediction of clinical efficacy of MSCT in the treatment of RA.4.Through a preclinical and clinical study to evaluate the regulatory capacity of human umbilical cord mesenchymal stem cells combined with recombinant human IFN-?on immune imbalance and inflammatory response in patients with RA,as well as the protective and repairing effects,and the efficacy and safety evaluation data of RA,,so as to explore new effective therapies for RA patients.Methods1.qPCR,WB and flow cytometry were used to detect the effects of TNF-?,IFN-?and TNF-?combined with IFN-?on the expression of PD-L1 and related molecular pathways.The molecular mechanism of TNF-?and IFN-?up-regulating the expression of PD-L1 in mesenchymal stem cells at translation level was also discussed in combination with specific inhibitors.2.We enrolled 15 severe sepsis patients who averagely divided into low?1×10⁶cells/kg?,intermediate?2×10⁶ cells/kg?,and high?3×10⁶ cells/kg?dosing cohort.Primary outcomes included the incidence of infusion associated events and serious adverse events.Secondary outcomes included systemic endpoints,mortality,and inflammation biologic markers.A historical case-matched comparison group was set as control.3.A total of 105 patients with persistently active RA and poor responses to traditional medication were randomly divided into MSCT and control groups.Outcomes were evaluated according to the 28-joint Disease Activity Score?DAS28?and Health Assessment Questionnaire?HAQ?,serological indicators,regulatory T cell?Treg?to T helper 17?Th17?cell ratio,and inflammatory cytokine levels.Twelve weeks after MSCT,the outcomes of the MSCT group were evaluated according to the European League against Rheumatism?EULAR?response criteria.Patients with a good or moderate response were added to the response group,and those with no response were added to the no-response group.4.mBMMSCs and IFN-?R^-/-mBMMSCs were transplanted into CIA mice by tail vein injection.The severity of arthritis was evaluated by clinical arthritis scoring system,histopathology and imaging methods.Sixty patients with active RA were randomly divided into two groups:mesenchymal stem cells?MSCs?group and mesenchymal stem cells combined with IFN-?treatment group?MSCs+IFN-??.Outcomes were evaluated according to activity score of 28 kinds of joint diseases?DAS28?and health assessment questionnaire?HAQ?,serological indicators,the ratio of regulatory T cells?Treg?to T helper cells 17?Th17?and the level of inflammatory cytokines.After 12 weeks of MSCT treatment,the efficacy of each group was evaluated according to EULAR response criteria,and patients with good or moderate response were defined as effective,patients without response were defined as ineffective.The difference of efficiency between the two groups was compared,and disease activity was further assessed by clustering.Results1.Our study found that TNF-?activates eIF4E by activating translation-related mTORC1-S6K and Ras-MAPK signaling pathways,and simultaneously increases the formation of active eukaryotic translation initiation complex eIF4F and IFN-?-induced MSCs PD-L1 mRNA levels in polysomes and significantly upgrades IFN-?-induced mesenchymal stem cells PD-L1 at translation level through post-transcriptional regulation,which enhances the immune regulatory capacity of MSCs.2.This study enrolled 15 severe sepsis patients?10 male and 5 female?with a median age of 58.There were no infusion associated serious events or treatment-related adverse events in any of the 15 patients in this trial,nor any safety or efficacy signals for serious adverse events or the measured cytokines between the interventional as compared to the historical case-matched comparison group.3.No serious adverse events were reported for either MSCT subgroup?28 in the response group and 24 in the no-response group?.The therapeutic effects lasted for 48weeks without continuous administration.Notably,a transient increase in serum IFN-???29?2pg/ml?levels was observed in the response group,but not in the no-response group.Furthermore,an increase in IL-10 levels and the Treg/Th17 ratio and a reduction in IL-6levels appeared 2-3 weeks after the transient IFN-?increase.4.Animal experiments showed that WT-MSCs could significantly improve the clinical severity of CIA,while IFN-?R^-/-MSCs could significantly aggravate synovitis,joint damage and cartilage damage in CIA mice.Compared with MSCs alone,the combined treatment of MSCs and IFN-?can significantly improve the clinical symptoms and disease activity of RA patients,improve the efficiency and improve the disease activity classification.Conclusion1.TNF-?significantly up-regulates IFN-?-induced mesenchymal stem cell PD-L1 at the translational level by post-transcriptional regulation,enhancing the immune regulation capacity of mesenchymal stem cells.2.A single intravenous infusion of allogeneic MSCs was safe and well tolerated in 15patients with severe sepsis up to a dose of 3×10⁶ cells/kg.3.Allogeneic MSCT is safe and feasible,and we propose high serum IFN-?levels as a potent biomarker for patient including and predicting MSCT response.4.This study combined with preclinical studies and clinical studies to demonstrate for the first time that IFN-?is a key factor in determining the efficacy of MSCs transplantation in the treatment of RA,and for the first time proposed a new clinical strategy for the treatment of RA with MSCs+IFN-?,and confirmed that this strategy can greatly improve the clinical efficacy of MSCs transplantation in the treatment of RA,with an effective rate of 93.3%.