Genomic alterations in human umbilical cord–derived mesenchymal stromal cells call for stringent quality control before any possible therapeutic approach

Background aimsThe umbilical cord (UC) is a promising source of mesenchymal stromal cells (MSCs). UC-MSCs display very similar in vitro characteristics to bone marrow–MSCs and could represent a valuable alternative for cell-based therapies. However, it is still unclear whether UC-MSCs are prone or not to the acquisition of genomic imbalances during in vitro expansion.MethodsWith the use of array-comparative genomic hybridization, we compared copy number variations of early (P2–P3) and late (>P5) passages of in vitro–expanded UC-MSCs.ResultsIn two of 11 long-term UC-MSCs cultures, we observed the appearance of clones carrying genomic imbalances, which generated genetic mosaicism at intermediate passages. Although still able to reach the senescence phase, the cells carrying the genomic imbalance acquired a proliferative advantage, as demonstrated by the increase in frequency during long-term culture.ConclusionsAltogether, our results suggest that UC-MSC–based clinical protocols should be designed with caution; their clinical use should be preceded by array-comparative genomic hybridization screening for the acquisition of genomic imbalances during in vitro expansion.     

Kanglaite injection plus platinum-based chemotherapy for stage III/IV non-small cell lung cancer: A meta-analysis of 27 RCTs

BackgroundKanglaite injection (KLT) is a broad-spectrum anti-tumor drug, which is extracted from the seeds of the Chinese medicinal herb Coix lacryma-jobi, and has been widely used for the treatment of advanced lung cancer.PurposeTo evaluate the combined effects of Kanglaite injection plus platinum-based chemotherapy (PBC) on patients with stage III/IV non-small cell lung cancer (NSCLC).Study designA systematic review and meta-analysis of randomized clinical trials (RCTs).Materials and methodsTwelve databases were searched from their inceptions until July 05, 2019. All the RCTs comparing the efficacy and safety of Kanglaite injection plus PBC versus PBC alone were selected. Analyses were performed using Review Manager 5.3, Comprehensive Meta-Analysis 3.0 and Trial Sequential Analysis (TSA). Disease control rate (DCR) was defined as the primary endpoint, objective response rate (ORR), survival rate, quality of life (QOL), cellular immunity function, and toxicities were defined as the secondary endpoints.ResultsTwenty-seven RCTs recruiting 2,243 patients with stage III/IV NSCLC were included. The results showed that, compared with PBC alone, Kanglaite injection plus PBC improved DCR (RR = 1.20, 95% CI 1.15–1.26, p < 0.00001), ORR (RR = 1.45, 95% CI 1.31–1.60, p < 0.00001), 1-year survival rate (RR = 1.20, 95% CI 1.02–1.43, p = 0.03), QOL (RR = 1.32, 95% CI 1.25–1.40, p < 0.00001), CD4⁺ T cells (WMD = 4.86, 95% CI 4.00–5.73, p < 0.00001), CD4⁺/CD8⁺ ratio (WMD = 0.19, 95% CI 0.07–0.31, p < 0.002), and reduced severe toxicities by 59% (RR = 0.41, 95% CI 0.33–0.51, p < 0.00001). Most results were robust and the quality of evidence was from moderate to low.ConclusionsKanglaite injection in combination with PBC showed significantly higher efficacy than PBC alone in the treatment of stage III/IV NSCLC. Moreover, the combination therapy can improve cellular immunity and attenuate the severe toxicities caused by chemotherapy. However, high-quality RCTs are warranted to further assess the effects of the combined therapy.     

Efficacy of granulocyte colony stimulating factor in patients with severe alcoholic hepatitis with partial or null response to steroid (GRACIAH trial): study protocol for a randomized controlled trial

Background

Alcoholic hepatitis (AH) has the most severe presentation among alcohol-related liver diseases. Corticosteroids are the most widely recommended treatment for severe AH. However, more innovative, refined treatment measures are required because of its high mortality despite corticosteroid treatment. This study aims to determine whether granulocyte colony stimulating factor (G-CSF) treatment increases short-term survival in patients with severe AH refractory to corticosteroid treatment.

Methods/design

Patients with severe AH whose Maddrey’s discriminant function (MDF) score is ≥?32 and who will be treated with prednisolone (40?mg/day) for 1?week will be screened. Among them, 190 subjects with a partial response (PR) (Lille score 0.16–0.56), and 78 subjects with a null response (NR) (Lille score?≥?0.56) will be enrolled. Subjects with PR will be randomized to steroid plus placebo or steroid plus 12?G-CSF injections (5?μg/kg/day for 5?days followed by every 3?days) at a ratio of 1:1. Subjects with a NR will be randomized to the placebo or G-CSF group (1:1). Study subjects in the PR group will be treated with prednisolone for 28?days followed by dose tapering for an additional 2?weeks. The primary endpoint is the 2-month survival rate in the NR group and the 6-month survival rate in the PR group. Child–Turcotte–Pugh, model for end-stage liver disease score, and the change in the proportion of peripheral circulating CD34-positive cells will be analyzed as risk factors for mortality. Preliminary safety data for the initial 10 study subjects enrolled in the PR study will be assessed to determine whether the PR study would be continued, according to the G-CSF-mobilized, peripheral-blood stem cell donor assessment protocol of the National Marrow Donor Program.

Discussion

We hypothesized that G-CSF would prolong short-term survival of patients with severe AH refractory to corticosteroid treatment. This is a proof-of-concept trial designed to assess the efficacy of Lille-score-guided G-CSF treatment. This trial is also designed to identify a special subgroup in whom G-CSF rescue treatment would improve liver function and prolong survival.

Trial registration

ClinicalTrials.gov, NCT02442180. Prospectively registered on 13 May 2015.

     

Effect of transplantation route on stem cell migration to fibrotic liver of rats via cellular magnetic resonance imaging

Background aimsAssessing mesenchymal stromal cells (MSCs) after grafting is essential for understanding their migration and differentiation processes. The present study sought to evaluate via cellular magnetic resonance imaging (MRI) if transplantation route may have an effect on MSCs engrafting to fibrotic liver of rats.MethodsRat MSCs were prepared, labeled with superparamagnetic iron oxide and scanned with MRI. Labeled MSCs were transplanted via the portal vein or vena caudalis to rats with hepatic fibrosis. MRI was performed in vitro before and after transplantation. Histologic examination was performed. MRI scan and imaging parameter optimization in vitro and migration under in vivo conditions were demonstrated.ResultsStrong MRI susceptibility effects could be found on gradient echo-weighted, or T21-weighted, imaging sequences from 24 h after labeling to passage 4 of labeled MSCs in vitro. In vivo, MRI findings of the portal vein group indicated lower signal in liver on single shot fast spin echo-weighted, or T2-weighted, imaging and T21-weighted imaging sequences. The low liver MRI signal increased gradually from 0–3 h and decreased gradually from 3 h to 14 days post-transplantation. The distribution pattern of labeled MSCs in liver histologic sections was identical to that of MRI signal. It was difficult to find MSCs in tissues near the portal area on day 14 after transplantation; labeled MSCs appeared in fibrous tuberculum at the edge of the liver. No MRI signal change and a positive histologic examination were observed in the vena caudalis group.ConclusionsThe portal vein route seemed to be more beneficial than the vena caudalis on MSC migration to fibrotic liver of rats via MRI.     

Effect of bone marrow–derived mesenchymal stromal cells on hepatoma

Background aimsThe aim of the study was to evaluate the effect of mesenchymal stromal cells (MSCs) on tumor cell growth in vitro and in vivo and to elucidate the apoptotic and anti-proliferative mechanisms of MSCs on a hepatocellular carcinoma (HCC) murine model.MethodsThe growth-inhibitory effect of MSCs on the Hepa 1–6 cell line was tested by means of methyl thiazolyl diphenyl-tetrazolium assay. Eighty female mice were randomized into four groups: group 1 consisted of 20 mice that received MSCs only by intrahepatic injection; group 2 consisted of 20 HCC mice induced by inoculation of Hepa 1–6 cells into livers without MSC treatment; group 3 consisted of 20 mice that received MSCs after induction of liver cancer; group 4 consisted of 20 mice that received MSCs after induction of liver cancer on top of induced biliary cirrhosis.ResultsMSCs exhibited a growth-inhibitory effect on Hepa 1–6 murine cell line in vitro. Concerning in vivo study, decreases of serum alanine transaminase, aspartate transaminase and albumin levels after MSC transplantation in groups 2 and 3 were found. Gene expression of α-fetoprotein was significantly downregulated after MSC injection in the HCC groups. We found that gene expression of caspase 3, P21 and P53 was significantly upregulated, whereas gene expression of Bcl-2 and survivin was downregulated in the HCC groups after MSC injection. Liver specimens of the HCC groups confirmed the presence of dysplasia. The histopathological picture was improved after administration of MSCs to groups 2 and 3.ConclusionsMSCs upregulated genes that help apoptosis and downregulated genes that reduce apoptosis. Therefore, MSCs could inhibit cell division of HCC and potentiate their death.     

Intrathoracic infusion therapy with Lentinan and chemical irritants for malignant pleural effusion: a systematic review and meta-analysis of 65 randomized controlled trials

BackgroundAderivative of Shiitake mushrooms, Lentinan is used to control malignant pleural effusion (MPE) through intrathoracic infusion.PurposeTo determine the clinical response, survival and safety of Lentinan plus chemical irritants, and the optimal combinations with chemical irritants, indication, threshold and optimal regimen for achieving the desired responses.Study designWe performed a new systematic review and meta-analysis following the PRISMA guidelines.MethodsWe collected all randomized controlled trials (RCTs) regarding Lentinan plus chemical irritants from Chinese and English electronic databases (from inception until March 2019). We evaluated their bias risk, synthesized data using meta-analysis, and summarized evidence quality following the Grades of Recommendation Assessment, Development and Evaluation approach.ResultsWe included 65 RCTs involving 4,080 patients and nine chemical irritants. Most trials had unclear bias risk. Lentinan with cisplatin significantly improved complete response [Risk ratio (RR) = 1.68, 95% confidence intervals (CI) (1.51 to 1.87), p < 0.00001, Fig.3a] and quality of life [RR = 1.51 95% CI (1.41 to 1.62), p < 0.00001, Fig.4], and decreased the risk of treatment failure, myelosuppression, gastrointestinal reaction, and chest pain. For patients with moderate to large volume of the pleural effusion, primary treatment, KPS score ≥ 50–60, or anticipated survival time ≥ 3months, Lentinan (3–4 mg/time, once a week for three to four times) withcisplatin (30–40 mg/m² or 50–60 mg/m²) significantly improved complete response and decreased failure. Most results were robust and moderate quality.ConclusionThe results suggest that Lentinan with chemical irritants, especially cisplatin is beneficial to the patient with MPE, and provide evidence for the indication, threshold, and optimal regimen that may achieve success and decrease failure.     

A chemically defined biomimetic surface for enhanced isolation efficiency of high-quality human mesenchymal stromal cells under xenogeneic/serum-free conditions

Background aimsMesenchymal stromal cells (MSCs) are one of the most frequently used cell types in regenerative medicine and cell therapy. Generating sufficient cell numbers for MSC-based therapies is constrained by (i) their low abundance in tissues of origin, which imposes the need for significant ex vivo cell expansion; (ii) donor-specific characteristics, including MSC frequency/quality, that decline with disease state and increasing age; and (iii) cellular senescence, which is promoted by extensive cell expansion and results in decreased therapeutic functionality. The final yield of a manufacturing process is therefore primarily determined by the applied isolation procedure and its efficiency in isolating therapeutically active cells from donor tissue. To date, MSCs are predominantly isolated using media supplemented with either serum or its derivatives, which poses safety and consistency issues.MethodsTo overcome these limitations while enabling robust MSC production with constant high yield and quality, the authors developed a chemically defined biomimetic surface coating called isoMATRIX (denovoMATRIX GmbH, Dresden, Germany) and tested its performance during isolation of MSCs.ResultsThe isoMATRIX facilitates the isolation of significantly higher numbers of MSCs in xenogeneic (xeno)/serum-free and chemically defined conditions. The isolated cells display a smaller cell size and higher proliferation rate than those derived from a serum-containing isolation procedure and a strong immunomodulatory capacity. The high proliferation rates can be maintained up to 5 passages after isolation and cells even benefit from a switch towards a proliferation-specific MSC matrix (myMATRIX MSC) (denovoMATRIX GmbH, Dresden, Germany).ConclusionIn sum, isoMATRIX promotes enhanced xeno/serum-free and chemically defined isolation of human MSCs and supports consistent and reliable cell performance for improved stem cell-based therapies.     

How important is differentiation in the therapeutic effect of mesenchymal stromal cells in liver disease?

Background aimsThe protocols for differentiation of hepatocyte-like cells (HLCs) from mesenchymal stromal cells (MSCs) have been well established. Previous data have shown that MSCs and their derived HLCs were able to engraft injured liver and alleviate injuries induced by carbon tetrachloride. The goal of the current study was to determine the differences of MSCs and their derived HLCs in terms of therapeutic functions in liver diseases.MethodsAfter hepatic differentiation of umbilical cord–derived MSCs in vitro, we detected both MSC and HLC expressions of adhesion molecules and chemokine receptor CXCR4 by flow cytometry; immunosuppressive potential and hepatocyte growth factor expression were determined by means of enzyme-linked immunosorbent assay. We compared the therapeutic effect for fulminant hepatic failure in a mouse model.ResultsMSC-derived-HLCs expressed lower levels of hepatocyte growth factor, accompanied by impaired immunosuppression in comparison with MSCs. Furthermore, undifferentiated MSCs showed rescuing potentials superior to those in HLCs for the treatment of fulminant hepatic failure.ConclusionsAfter differentiation, HLCs lost several major properties in comparison with undifferentiated MSCs, which are beneficial for their application in liver diseases. Undifferentiated MSCs may be more appropriate than are HLCs for the treatment of liver diseases.     

Mesenchymal stromal cell injection protects against oxidative stress in Escherichia coli–induced acute lung injury in mice

Background aimsStem cells may be a promising therapy for acute respiratory distress syndrome. Recent in vivo and in vitro studies suggested that the mesenchymal stromal cells (MSCs) have anti-oxidative stress properties. We hypothesized that intravenous injection of bone marrow–derived mesenchymal stem cells (MSCs) could attenuate Escherichia coli–induced acute lung injury (ALI) in mice by controlling the oxidative stress status.MethodsEighty mice were randomly divided into four groups: group 1 (control group) received 25 μL of saline as a vehicle; group 2 contained E coli–induced ALI mice; group 3 included mice that received MSCs before induction of ALI; group 4 included mice that received MSCs after induction of ALI. Lung samples were isolated and assayed for oxidative stress variables and histopathologic analysis. Total anti-oxidant capacity was measured in broncho-alveolar lavage.ResultsPre- and post-injury MSC injection increased survival, reduced pulmonary edema and attenuated lung injuries in ALI mice. Histologically, MSCs exhibited a considerable degree of preservation of the pulmonary alveolar architecture. An increase of anti-oxidant enzyme activities and a decrease of myeloperoxidase activity and malondialdehyde levels in the MSC recipient groups versus the ALI group were found. Furthermore, the total anti-oxidant capacity and reduced glutathione levels were significantly increased in MSCs recipient groups versus the ALI group. Weak +ve inducible nitric oxide synthase immuno-expression in groups that received MSCs was detected. Pre-injury MSC injection showed better effects than did post-injury MSC injection.ConclusionsSystemic bone marrow–derived MSC injection was effective in modulating the oxidative stress status in E coli–induced acute lung injury in mice.     

Optimization of the therapeutic efficacy of human umbilical cord blood–mesenchymal stromal cells in an NSG mouse xenograft model of graft-versus-host disease

Background aimsAlthough in vitro studies have demonstrated the immunosuppressive capacity of mesenchymal stromal cells (MSCs), most in vivo studies on graft-versus-host disease (GVHD) have focused on prevention, and the therapeutic effect of MSCs is controversial. Moreover, optimal time intervals for infusing MSCs have not been established.MethodsWe attempted to evaluate whether human umbilical cord blood–MSCs (hUCB-MSCs) could either prevent or treat GVHD in an NSG mouse xenograft model by injection of MSCs before or after in vivo clearance. Mice were infused with either a single dose or multiple doses of 5 × 10⁵ hUCB-MSCs (3- or 7-day intervals) before or after GVHD onset.ResultsBefore onset, hUCB-MSCs significantly improved the survival rate only when repeatedly injected at 3-day intervals. In contrast, single or repeated injections after GVHD onset significantly increased the survival rate and effectively attenuated tissue damage and inflammation. Furthermore, the levels of prostaglandin E₂ and transforming growth factor-β1 increased significantly, whereas the level of interferon-γ decreased significantly in all MSC treatment groups.ConclusionsThese data establish the optimal time intervals for preventing GVHD and show that hUCB-MSCs effectively attenuated symptoms and improved survival rate when administered after the onset of GVDH.     

Proceedings of the ISCT scientific signature series symposium, “Advances in cell and gene therapies for lung diseases and critical illnesses”: International Society for Cell & Gene Therapy,Burlington VT,US, July 16, 2021

The ISCT Scientific Signature Series Symposium “Advances in Cell and Gene Therapies for Lung Diseases and Critical Illnesses” was held as an independent symposium in conjunction with the biennial meeting, “Stem Cells, Cell Therapies, and Bioengineering in Lung Biology and Diseases,” which took place July 12–15, 2021, at the University of Vermont. This is the third Respiratory System–based Signature Series event; the first 2, “Tracheal Bioengineering, the Next Steps” and “Cellular Therapies for Pulmonary Diseases and Critical Illnesses: State of the Art of European Science,” took place in 2014 and 2015, respectively. Cell- and gene-based therapies for respiratory diseases and critical illnesses continue to be a source of great promise and opportunity. This reflects ongoing advancements in understanding of the mechanisms by which cell-based therapies, particularly those using mesenchymal stromal cells (MSCs), can mitigate different lung injuries and the increasing sophistication with which preclinical data is translated into clinical investigations. This also reflects continuing evolution in gene transfer vectors, including those designed for in situ gene editing in parallel with those targeting gene or cell replacement. Therefore, this symposium convened global thought leaders in a forum designed to catalyze communication and collaboration to bring the greatest possible innovation and value of cell- and gene-based therapies for patients with respiratory diseases and critical illnesses.     

Clinical trials with haemopoietic growth factors

Five glycoprotein growth factors capable of stimulating the proliferation and differentiation of haemopoietic progenitor cells in vitro have been identified and sequenced over the past ten years. Recombinant DNA technology has recently enabled the production of sufficient amounts of these agents for preclinical testing. Erythropoietin (EPO), granulocyte-macrophage colony-stimulating factor (GM-CSF), and granulocyte colony-stimulating factor (G-CSF) have already entered clinical studies in humans. Interleukin-3 (IL-3) and macrophage colony-stimulating factor (M-CSF) should soon be available for use in humans. EPO corrects the anaemia of end stage renal failure, improving the quality of life for such patients and preventing the need for red cell transfusions. At high dose it increases platelet production in vitro and in vivo and may be of value in humans to prevent the thrombocytopaenia associated with chemotherapy. G-CSF and GM-CSF have been used in several clinical studies. Administration of both growth factors results in a leucocytosis, G-CSF predominantly increasing neutrophil production and GM-CSF increasing production of neutrophils, eosinophils and monocytes. The optimal administration of these agents is via continuous intravenous infusion or daily subcutaneous injections at doses of 3–10μg/kg/24 h. GM-CSF has shown promising results in patients with AIDS and the myelodysplastic syndrome and both G-CSF and GM-CSF have reduced the duration of neutropaenia and incidence of infection associated with chemotherapy. These agents may allow an escalation of the dose-intensity of chemotherapy in the future and thereby, hopefully, increase the response rate and survival for patients with a variety of neoplasms. Several other potential roles for these haemopoietic growth factor are discussed.     

Human mesenchymal stromal cells transiently increase cytokine production by activated T cells before suppressing T-cell proliferation: effect of interferon-γ and tumor necrosis factor-α stimulation

Background aimsMesenchymal stromal cells (MSCs) suppress T-cell proliferation, especially after activation with inflammatory cytokines. We compared the dynamic action of unprimed and interferon (IFN)-γ plus tumor necrosis factor (TNF)-α–pretreated human bone marrow–derived MSCs on resting or activated T cells.MethodsMSCs were co-cultured with allogeneic peripheral blood mononuclear cells (PBMCs) at high MSC-to-PBMC ratios in the absence or presence of concomitant CD3/CD28-induced T-cell activation. The kinetic effects of MSCs on cytokine production and T-cell proliferation, cell cycle and apoptosis were assessed.ResultsUnprimed MSCs increased the early production of IFN-γ and interleukin (IL)-2 by CD3/CD28-activated PBMCs before suppressing T-cell proliferation. In non-activated PBMC co-cultures, low levels of IL-2 and IL-10 synthesis were observed with MSCs in addition to low levels of CD69 expression by T cells and no T-cell proliferation. MSCs also decreased apoptosis in resting and activated T cells and inhibited the transition of these cells into the sub-G0/G1 and the S phases. With inhibition of indoleamine 2,3 dioxygenase, MSCs increased CD3/CD28-induced T-cell proliferation. After priming with IFN-γ plus TNF-α, MSCs were less potent at increasing cytokine production by CD3/CD28-activated PBMCs and more effective at inhibiting T-cell proliferation but had preserved anti-apoptotic functions.ConclusionsUnprimed MSCs induce a transient increase in IFN-γ and IL-2 synthesis by activated T cells. Pre-treatment of MSCs with IFN-γ plus TNF-α may increase their effectiveness and safety in vivo.     

Heterotopic bone formation derived from multipotent stromal cells is not inhibited in aged mice

Background aimsDecreased bone formation with age is believed to arise, at least in part, because of the influence of the senescent microenvironment. In this context, it is unclear whether multipotent stromal cell (MSC)-based therapies would be effective for the treatment of bone diseases.MethodsWith the use of a heterotopic bone formation model, we investigated whether MSC-derived osteogenesis is impaired in aged mice compared with young mice.ResultsWe found that bone formation derived from MSCs is not reduced in aged mice. These results are supported by the unexpected finding that conditioned media collected from ionizing radiation–induced senescent MSCs can stimulate mineralization and delay osteoclastogenesis in vitro.ConclusionsOverall, our results suggest that impaired bone formation with age is mainly cell-autonomous and provide a rationale for the use of MSC-based therapies for the treatment of bone diseases in the elderly.     

Analysis of chemotactic molecules in bone marrow-derived mesenchymal stem cells and the skin: Ccl27-Ccr10 axis as a basis for targeting to cutaneous tissues

Background aimsAdult stem cells produce a plethora of extracellular matrix molecules and have a high potential as cell-based therapeutics for connective tissue disorders of the skin. However, the primary challenge of the stem cell-based approach is associated with the inefficient homing of systemically infused stem cells to the skin.MethodsWe examined chemotactic mechanisms that govern directional migration of mesenchymal stem cells (MSCs) into the skin by conducting a comprehensive expression analysis of chemotactic molecules in MSCs and defined cutaneous tissues from normal and hereditary epidermolysis bullosa (EB)-affected skin.ResultsAnalysis of chemokine receptors in short-term and long-term MSC cultures showed tissue culture-dependent expression of several receptors. Assessment of epidermis-derived and dermis-derived chemokines showed that most chemotactic signals that originate from the skin preferentially recruit different sets of leukocytes rather than MSCs. Analysis of the chemotactic molecules derived from EB-affected non-blistered skin showed only minor changes in expression of selected chemokines and receptors. Nevertheless, the data allowed us to define the Ccl27-Ccr10 chemotactic axis as the most potent for the recruitment of MSCs to the skin. Our in vivo analysis demonstrated that uniform expression of Ccr10 on MSCs and alteration of Ccl27 level in the skin enhance extravasation of stem cells from circulation and facilitate their migration within cutaneous tissue.ConclusionsCollectively, our study provides a comprehensive analysis of chemotactic signals in normal and EB-affected skin and proof-of-concept data demonstrating that alteration of the chemotactic pathways can enhance skin homing of the therapeutic stem cells.     

G-CSF promotes the viability and angiogenesis of injured liver via direct effects on the liver cells

Background

Presently, liver transplantation is the only treatment strategy for liver failure (LF). Although granulocyte-colony stimulating factor (G-CSF) exhibits protective functions in LF, it is not clear whether it directly affects the liver cells.

Methods and Results

We established an injured liver cell model and observed that G-CSF treatment promoted cell viability and enhanced Ki67 and VEGF-A expression. Thereafter, human umbilical vein endothelial cells (HUVECs) were cultured in a conditioned medium collected from the G-CSF-treated injured liver cells. HUVECs’ proliferation and tubule formation were promoted. Furthermore, in an injured liver mouse model, confirmed via haematoxylin–eosin staining, we evaluated serum alanine aminotransferase activity, Ki67 expression, and microvessel density (MVD). G-CSF treatment significantly relieved liver injury, upregulated Ki67 expression, and enhanced MVD in the injured mouse liver tissue. Additionally, AKT and ERK signal targets were explored, and it was demonstrated that the effects of G-CSF on injured liver cells were mediated through the AKT and ERK signalling pathways.

Conclusions

G-CSF promotes injured liver viability and angiogenesis by directly affecting injured liver cells via the AKT and ERK signalling pathways. These findings improve our understanding of the role of G-CSF in recovery from LF.

     

Encapsulating Non-Human Primate Multipotent Stromal Cells in Alginate via High Voltage for Cell-Based Therapies and Cryopreservation

Alginate cell-based therapy requires further development focused on clinical application. To assess engraftment, risk of mutations and therapeutic benefit studies should be performed in an appropriate non-human primate model, such as the common marmoset (Callithrix jacchus). In this work we encapsulated amnion derived multipotent stromal cells (MSCs) from Callithrix jacchus in defined size alginate beads using a high voltage technique. Our results indicate that i) alginate-cell mixing procedure and cell concentration do not affect the diameter of alginate beads, ii) encapsulation of high cell numbers (up to 10×10⁶ cells/ml) can be performed in alginate beads utilizing high voltage and iii) high voltage (15–30 kV) does not alter the viability, proliferation and differentiation capacity of MSCs post-encapsulation compared with alginate encapsulated cells produced by the traditional air-flow method. The consistent results were obtained over the period of 7 days of encapsulated MSCs culture and after cryopreservation utilizing a slow cooling procedure (1 K/min). The results of this work show that high voltage encapsulation can further be maximized to develop cell-based therapies with alginate beads in a non-human primate model towards human application.     

Tumor stroma engraftment of gene-modified mesenchymal stem cells as anti-tumor therapy against ovarian cancer

Background aimsMany ovarian cancers originate from ovarian surface epithelium, where they develop from cysts intermixed with stroma. The stromal layer is critical to the progression and survival of the neoplasm and consequently is recruited into the tumor microenvironment.MethodsUsing both syngeneic mouse tumors (ID8-R) and human xenograft (OVCAR3, SKOV3) tumor models, we first confirmed that intraperitoneally injected circulating mesenchymal stem cells (MSCs) could target, preferentially engraft and differentiate into α-smooth muscle actin-positive myofibroblasts, suggesting their role as “reactive stroma” in ovarian carcinoma development and confirming their potential as a targeted delivery vehicle for the intratumoral production of interferon-β (IFN-β). Mice with ovarian carcinomas then received weekly intraperitoneal injections of IFN-β expressing MSCs.ResultsIntraperitoneal injections of IFN-β expressing MSCs resulted in complete eradication of tumors in 70% of treated OVCAR3 mice (P = 0.004) and an increased survival of treated SKOV3 mice compared with controls (P = 0.01). Similar tumor growth control was observed using murine IFN-β delivered by murine MSCs in ID8-R ovarian carcinoma. As a potential mechanism of tumor killing, MSCs produced IFN-β-induced caspase-dependent tumor cell apoptosis.ConclusionsOur results demonstrate that ovarian carcinoma engrafts MSCs to participate in myofibrovascular networks and that IFN-β produced by MSCs intratumorally modulates tumor kinetics, resulting in prolonged survival.