Allogeneic Bone Marrow-derived Mesenchymal Stem Cells Research Articles

CellKine clinical trial: first report from a phase 1 trial of allogeneic bone marrow–derived mesenchymal stem cells in subjects with painful lumbar facet joint arthropathy

Abstract Background: Lumbar facet joint arthropathy (LFJA) is a major cause of low back pain (LBP), with current treatments offering limited long-term benefits. Bone marrow–derived mesenchymal stem cells (BM-MSCs) show promise due to their immunomodulatory and trophic effects, potentially addressing underlying degenerative processes in LFJA. Objectives: This initial report describes the outcomes of the first treated patient in an ongoing mutidisciplinary phase 1 clinical trial evaluating the safety and feasibility of intra-articular allogeneic BM-MSCs for painful LFJA. Methods: Following enrollment in our IRB-approved protocol, symptomatic LFJA was confirmed through double blocks on L4 and L5 medial branches. Two 1-mL syringes, each containing 10 million BM-MSCs, were prepared in the cGMP facility and administered bilaterally to the patient’s L4-L5 lumbar facet joints. The patient underwent standardized follow-ups, including clinical examinations and functional and imaging assessments for 2 years, utilizing patient-reported outcomes measurement information system—computer adaptive tests (PROMIS CATs), visual analogue scale, Oswestry disability index, work functional status and opioid pain medication use, and MR imaging Fenton–Czervionke score. Results: The patient tolerated the procedure well, with no drug-related adverse events during the study period. Pain, spine function, and work functional status improved at multiple follow-ups. This patient also reported improvements in mental and social health, along with a notable improvement in the grade of facet synovitis observed at the one-year follow-up MRI evaluation. Conclusions: This case report suggests the safety and feasibility of administering intra-articular allogeneic BM-MSCs, offering therapeutic benefits for pain management and functional activities.

The treatment efficacy of bone tissue engineering strategy for repairing segmental bone defects under diabetic condition.

Diabetes mellitus is a systematic disease which exert detrimental effect on bone tissue. The repair and reconstruction of bone defects in diabetic patients still remain a major clinical challenge. This study aims to investigate the potential of bone tissue engineering approach to improve bone regeneration under diabetic condition. In the present study, decalcified bone matrix (DBM) scaffolds were seeded with allogenic fetal bone marrow-derived mesenchymal stem cells (BMSCs) and cultured in osteogenic induction medium to fabricate BMSC/DBM constructs. Then the BMSC/DBM constructs were implanted in both subcutaneous pouches and large femoral bone defects in diabetic (BMSC/DBM in DM group) and non-diabetic rats (BMSC/DBM in non-DM group), cell-free DBM scaffolds were implanted in diabetic rats to serve as the control group (DBM in DM group). X-ray, micro-CT and histological analyses were carried out to evaluate the bone regenerative potential of BMSC/DBM constructs under diabetic condition. In the rat subcutaneous implantation model, quantitative micro-CT analysis demonstrated that BMSC/DBM in DM group showed impaired bone regeneration activity compared with the BMSC/DBM in non-DM group (bone volume: 46 ± 4.4mm3 vs 58.9 ± 7.15mm3, *p < 0.05). In the rat femoral defect model, X-ray examination demonstrated that bone union was delayed in BMSC/DBM in DM group compared with BMSC/DBM in non-DM group. However, quantitative micro-CT analysis showed that after 6months of implantation, there was no significant difference in bone volume and bone density between the BMSC/DBM in DM group (199 ± 63mm3 and 593 ± 65mg HA/ccm) and the BMSC/DBM in non-DM group (211 ± 39mm3 and 608 ± 53mg HA/ccm). Our data suggested that BMSC/DBM constructs could repair large bone defects in diabetic rats, but with delayed healing process compared with non-diabetic rats. Our study suggest that biomaterial sacffolds seeded with allogenic fetal BMSCs represent a promising strategy to induce and improve bone regeneration under diabetic condition.

Proliferation and Differentiation Potential of Bone Marrow-Derived Mesenchymal Stem Cells From Children With Polydactyly and Adults With Basal Joint Arthritis.

This study compared the proliferation and differentiation potential of bone marrow-derived mesenchymal stem cells (BMSCs) derived from infants with polydactyly and adults with basal joint arthritis. The proliferation rate of adult and infant BMSCs was determined by the cell number changes and doubling times. The γH2AX immunofluorescence staining, age-related gene expression, senescence-associated β-galactosidase (SA-β-gal) staining were analyzed to determine the senescence state of adult and infant BMSCs. The expression levels of superoxide dismutases (SODs) and genes associated with various types of differentiation were measured using Real-Time Quantitative Polymerase Chain Reaction (RT-qPCR). Differentiation levels were evaluated through histochemical and immunohistochemical staining. The results showed that infant BMSCs had a significantly higher increase in cell numbers and faster doubling times compared with adult BMSCs. Infant BMSCs at late stages exhibited reduced γH2AX expression and SA-β-gal staining, indicating lower levels of senescence. The expression levels of senescence-related genes (p16, p21, and p53) in infant BMSCs were also lower than in adult BMSCs. In addition, infant BMSCs demonstrated higher antioxidative ability with elevated expression of SOD1, SOD2, and SOD3 compared with adult BMSCs. In terms of differentiation potential, infant BMSCs outperformed adult BMSCs in chondrogenesis, as indicated by higher expression levels of chondrogenic genes (SOX9, COL2, and COL10) and positive immunohistochemical staining. Moreover, differentiated cells derived from infant BMSCs exhibited significantly higher expression levels of osteogenic, tenogenic, hepatogenic, and neurogenic genes compared with those derived from adult BMSCs. Histochemical and immunofluorescence staining confirmed these findings. However, adult BMSCs showed lower adipogenic differentiation potential compared with infant BMSCs. Overall, infant BMSCs demonstrated superior characteristics, including higher proliferation rates, enhanced antioxidative activity, and greater differentiation potential into various lineages. They also exhibited reduced cellular senescence. These findings, within the context of cellular differentiation, suggest potential implications for the use of allogeneic BMSC transplantation, emphasizing the need for further in vivo investigation.

Intravitreal allogeneic mesenchymal stem cells: a non-randomized phase II clinical trial for acute non-arteritic optic neuropathy

BackgroundAn effective treatment for acute non-arteritic ischemic optic neuropathy (NA-AION) has not been known or proven yet. Previous studies have suggested a neuroprotective effect of allogeneic bone marrow-derived mesenchymal stem cells. This study aims to report the results of a clinical trial on patients with acute non-arteritic optic neuropathy (NA-AION) treated with an intravitreal injection of allogeneic bone marrow-derived mesenchymal stem cells (BM-MSCs) (MSV®).MethodsWe conducted a prospective, non-randomized, clinical phase-II study (Eudra CT number 2016-003029-40; ClinicalTrials.gov Registry NCT03173638) that included 5 patients with acute unilateral NA-AION diagnosed within 2 weeks after symptom onset and who received an intravitreal injection of allogeneic BM-MSCs (0.05 ml; cell concentration: 1.5 × 106cells/mL). The patients underwent regular ophthalmological examinations and were followed for one year.ResultsIn this trial, allogeneic BM-MSCs appeared to be safe as no patients developed signs of acute nor chronic intraocular inflammation or a significant change in intraocular pressure, although an epiretinal membrane was developed in one patient. A retrolental aggregate formed shortly after the injection spontaneously disappeared within a few weeks in another phakic patient, leaving a subcapsular cataract. Visual improvement was noted in 4 patients, and amplitudes of P100 on the visually evoked potentials recordings increased in three patients. The retinal nerve fiber layer and macular ganglion cell layer thicknesses significantly decreased during the follow-up.ConclusionsBesides the development of an epiretinal membrane in one patient, the intravitreal application of allogeneic BM-MSCs appeared to be intraocularly well tolerated. Consequently, not only NA-AION but also BM-MSCs deserve more clinical investigational resources and a larger randomized multicenter trial that would provide stronger evidence both about safety and the potential therapeutic efficacy of intravitreally injected allogeneic BM-MSCs in acute NA-AION.Trial registration: Safety Assessment of Intravitreal Mesenchymal Stem Cells for Acute Non-Arteritic Anterior Ischemic Optic Neuropathy (NEUROSTEM). NCT03173638. Registered June 02, 2017 https://clinicaltrials.gov/ct2/show/NCT03173638.

Treatment of chronic liver disease using cell‑engineered constructs: morphofunctional characteristics

Objective: to study the effectiveness of correcting the morphofunctional characteristics of the liver in an experimental model of chronic liver disease (CLD), using implanted cell-engineered constructs (CECs).Materials and methods. Experiments were carried out on male Wistar rats (n = 80) aged 6–8 months with an initial weight of 230–250 g. CLD was modeled by inoculating the rats with 60% CCl4 oil solution for 42 days based on a modified scheme. Microgel based on recombinant spidroin rS1/9 was used as a matrix for CECs fabrication. Allogeneic liver cells (LCs) and multipotent bone marrow-derived mesenchymal stem cells (BM-MSCs) from a healthy donor were used as the cellular component of the CECs. The effectiveness of the corrective effect of the implanted CECs was assessed in an experimental CLD model (n = 60) in two groups of rats: Group 1 (control, n = 20, 1 mL of saline solution was injected into the damaged liver parenchyma) and Group 2 (experimental, n = 40, CECs containing allogenic LCs and BM-MSCs in a 5 : 1 ratio in a volume of 1 mL were implanted into the damaged liver parenchyma). For long-term monitoring of the CEC state, the CECs were labeled by additional inclusion in Cytodex-3. The effectiveness of the regulatory effect of CECs on regenerative processes in the liver was evaluated using biochemical, morphological and morphometric techniques, as well as by flow cytometry at 90 days after implantation.Results. In the control group, the mortality rate in CLD was 25%. There was no death in the experimental group with CLD after CEC implantation. The CECs were found to have a corrective effect on the biochemical and morphological parameters of the liver in CLD during 90 days of follow-up, with concomitant preservation of structural cellular homeostasis in the implanted CECs. Conclusion. Implantation of CECs in the liver facilitates effective correction of CLD by activating regenerative processes in the damaged liver, which is due to long-term preservation of structural cellular homeostasis in the CECs.

Tratamiento de la laminitis crónica en equinos utilizando células troncales mesenquimales alogénicas de la médula ósea

Chronic laminitis is a disabling condition that affects the laminar corium of the horse’s hooves. Commonly, it develops as a collateral injury of numerous primary systemic diseases. It is believed that the critical physiopathological event that renders a hoof laminitic is the loss of mesenchymal stem cells. This loss greatly impairs the ability of the laminar corium to regenerate. Although previous work provides credibility to this notion, there remain unsettled issues that must be addressed before accepting it as a well-founded fact. Here, it was reexamined the central tenet of the physiopathological model of laminitis by infusing allogeneic bone marrow-derived mesenchymal stem cells (ABM-MSCs), through the digital palmar vein, into the hooves of horses afflicted by chronic laminitis. Horses were clinically monitored during 6 mo by evaluating them monthly using the lameness-modified Obel-Glasgow’s scale and hooves thermography. Venograms and lamellar biopsies were taken at the beginning and at the end of the study period to gathered evidence on vascular remodeling and laminar corium regeneration. The results showed that ABM-MSCs infusion promotes vascular remodeling and laminar corium regeneration, further supporting that the loss of stem cells is the critical event leading to chronic laminitis. This work also demonstrated that the infusion of ABM-MSCs is safe since the treated horses did not develop local or systemic, negative clinical manifestations attuned with rejection reactions, at least during the 6-mo period they were follow up and under the therapeutic scheme proposed.

Safety and feasibility of subconjunctival injection of mesenchymal stem cells for acute severe ocular burns: A single-arm study

PurposeTo investigate the safety and feasibility of topical injection of bone marrow derived mesenchymal stem cells (BM-MSCs) in acute severe ocular burns. MethodsIn this open-label，single-arm study, subconjunctival injection of allogenic BM-MSCs combined with standard treatment was given to 16 patients with acute severe ocular burns (Dua's grade IV to VI) within 2 weeks after injury. The primary outcome was efficacy rate which referred to the proportion of complete corneal epithelialization patients without perforation. The secondary outcome was safety, visual acuity, the number of symblephara, and elevated intraocular pressure. ResultsOne patient was lost to follow-up. During the follow-up period, no corneal perforation was developed. Complete corneal epithelialization was noted 8 (ranged 4–10 weeks) weeks after treatment in 13 eyes (81.3%). The efficacy rate was 87.5% (95% confidence interval, CI: 61.7–98.4). Hypopyon was detected and later well controlled in 1 eye. Partial or total limbal stem cell deficiency (LSCD) was noted in all eyes. Improvement of visual acuity was achieved in 5 out of 16 eyes (31.3%). Seven eyes’ visual acuity was reached 0.1. Symblepharon with varied severity was noted in 5 eyes. Two eyes had elevated intraocular pressure. ConclusionsThis study confirms the safety of subconjunctival injection of BM-MSCs as an innovative and convenient procedure in ocular burns. The overall result is promising considering the absence of perforation, the low severity of symblepharon and visual acuity improvement.

Do allogeneic bone marrow derived mesenchymal stem cells diminish the inflammatory response to lipopolysaccharide infusion in horses? A pilot study

Endotoxemia is a leading cause of morbidity and mortality in the equine industry, with colic being the most common cause of endotoxemia in horses. The objective of this study was to evaluate the safety and potential efficacy of a single dose of allogeneic equine bone marrow derived mesenchymal stem cells (BM-MSCs) in horses after the IV administration of lipopolysaccharide (LPS).Six horses were administered an IV infusion of 30 ng/kg LPS (O55:B5 Escherichia coli) in 500 ml saline over 30 min. Immediately after infusion test horses (n = 3) were administered 100 × 106 allogeneic BM-MSCs diluted in saline IV and control horses (n = 3) were administered saline. Clinicopathological data, pro-inflammatory cytokine measurements and sCD14 concentrations were compared between groups.No adverse reactions were observed in horses administered BM-MSCs intravenously. There were no significant differences between test and control horses with regard to clinicopathological values or pro-inflammatory cytokine production. At no time point did concentrations of sCD14 exceed the reference range in any horse.Results suggest that administration of a single IV dose of freshly cultured MSCs is safe and well-tolerated in horses with induced endotoxemia. Further study to evaluate their efficacy as a potential therapeutic in a larger number of horses with clinical disease is required.

Allogeneic Bone Marrow Mesenchymal Stem Cell Transplantation in Tooth Extractions Sites Ameliorates the Incidence of Osteonecrotic Jaw-Like Lesions in Zoledronic Acid-Treated Rats.

Medication-related osteonecrosis of the jaw (MRONJ) is defined as the exposed necrotic bone involving the maxillofacial structures in bisphosphonate treated patients, and the pathophysiology of this disease remains unclear. The aim of this study was to assess the effects of the allogeneic transplantation of bone marrow-derived mesenchymal stem cells (BM-MSCs) in a model of Wistar mice with induced MRONJ disease. BM-MSCs from five male Wistar rats were characterized and cultured on β-tricalcium phosphate (β-TCP) granules. Thirty female Wistar rats were injected intraperitoneally with zoledronic acid and afterwards upper jaw molars were extracted. The animals were randomized to receive: Group 1: 1 × 106 BM-MSCs/β-TCP construct in the alveolar socket; and Group 2: Saline solution/β-TCP construct. A clinical and histological analysis was performed. Nested polymerase chain reaction (PCR) was assessed to verify the presence of transplanted male rat cells in the female recipient jaws. Clinical and histological findings evidenced that none of the animals in Group 1 exhibited uncovered sockets or bone exposure associated to MRONJ, whereas we detected 33% of MRONJ cases in Group 2. In addition, male rat cells were detected in the maxillae site four weeks after transplantation in the BM-MSCs-group. Allogeneic BM-MSCs in extractions sites ameliorates MRONJ incidence in zoledronic acid-treated rats compared to non-MSC treatments.

Allogeneic bone marrow derived mesenchymal stem cells outperform the autologous in modulating dendritic cells isolated from type 1 diabetic patients

Background & Aim Dendritic cells (DCs) have been implicated in the pathogenesis of type 1 diabetes mellitus (T1DM) and have potential as immunotherapeutic agents. The inhibition of DCs profile and function is part of the multi-level immunomodulatory properties displayed by human Mesenchymal Stem Cells (MSCs). Due to their potent immunoregulatory and immunomodulatory functions, MSCs have emerged as a therapeutic regimen in the management of many diseases that have no effective treatment, including T1DM. It has been shown that in T1DM patients DCs respond in terms of maturation, activation, and phenotype when co-cultured with autologous and allogeneic Bone Marrow derived MSCs (BM-MSC). In the recent study, the effect of autologous BM-MSCs in comparison with the allogeneic BM-MSCs co-cultured with monocyte-derived DCs isolated from TIDM patients was investigated for the first time. Methods, Results & Conclusion Methods Autologous and allogeneic BM-MSCs were isolated from healthy donors and T1DM patients. Then, morphology and surface markers expression were analyzed. Mature DCs (mDCs) were differentiated from CD14+ cells, obtained from newly diagnosed T1DM patients, pulsed with antigen Glutamic Acid Decarboxylase (GAD65) and co-cultured with BM-MSCs. the expression of co-stimulatory molecules and maturation markers were evaluated by flow cytometery. The level of anti-inflammatory cytokines was measured by ELISA. Autologous CD14- cells and GAD65 pulsed mDCs were co-cultured and IFN-γ ELISPOT responses were performed. Secreted level of IL-17 was measured Result autologous and allogeneic BM-MSCs have a comparable effect on GAD65 pulsed mature mDCs in terms of phenotype and maturation marker expression. Interestingly, the secretion levels of (IL-10, IL-6 and TGF-B) increased significantly in the allogeneic BM-MSCs when co-cultured with GAD65 pulsed mDCs. GAD65 pulsed-mDCs/T-cells showed decreased IFN-γ production when co-cultured with BM-MSCs. Moreover, the level of IL-17 secretion declined significantly in the allogeneic BM-MSCs compared to the autologous cells. Conclusion This research demonstrated in vitro the potent immunomodulatory effect of allogeneic BM-MSCs on pancreatic islet antigen specific T-Cells in newly-diagnosed T1DM patients through the induction of DCs’ tolerogenic function. These DCs can contribute to the suppressing T-cell mediated immunity and the promotion of anti-inflammatory responses exerted by BM-MSCs.

The MODULATORY EFFECTS OF BONE MARROW-DERIVED MESENCHMAL STEM CELLS ON CYCLOPHOSPHAMIDE INDUCED HEPATOTOXICITY IN CARCINOMA MICE

Objective: Cyclophosphamide (CPA) is a chemotherapeutic agent, induces hepatotoxicity as one of its side effects. Therefore, the present work was designed to investigate the protective role of bone marrow-derived mesenchymal stem cells (BM-MSCs) on CPA–induced hepatotoxicity in Ehrlich Ascites Carcinoma bearing-mice (EAC) and to test whether BM-MSCs influences the antitumor properties of the CPA.&#x0D; Methods: The hepatoprotective effects of BM-MSCs (single dose of 100 µl of a cell suspension containing allogenic BM-MSCs, i. v.) was evaluated in a model of hepatotoxicity by CPA (10 mg/kg/d i. p.) in EAC-female mice for one month. The anti-tumor activities of CPA and BM-MSCs were assessed by measuring mean tumor weight, mean survival time and the increase in life span. Moreover, ALT, AST, GGT, MDA, GSH, SOD, IL-6, IL10, caspase-3 and Bcl2 were measured.&#x0D; Results: The i. p. administration of CPA and BM-MSCs resulted in significant reductions in tumor size and mean tumor weight as well as caused concurrent significant increases in the life span as compared to the EAC mice. Furthermore, BM-MSCs ameliorated the liver enzyme markers namely ALT, AST, GGT, and hepatic oxidative stress through inhibition of MDA level that correlated with significant improvement in antioxidant status via increasing GSH and SOD levels as compared to both EAC and EAC+CPA groups. Moreover, BM-MSCs treatment significantly reduced the inflammatory marker level IL-6 as well as increment the level of IL-10 with subsequent decreases apoptosis via a depletion in the caspase-3 associated with an enhancement in the level of Bcl2 as compared to EAC group and EAC+CPA group. Minor histological lesions were observed in the liver tissue sections of mice treated with CPA and BM-MSCs as compared to the high histological lesions observed in the liver of the EAC group and CPA treated group.&#x0D; Conclusion: These results concluded that the combination treatment of BM-MSCs with CPA exhibited promising potential antitumor efficacy with greater safety than CPA treatment alone in mice via its antioxidant, anti-inflammatory and antiapoptotic effects.

Therapeutic Potential of Allogenic Bone Marrow-Derived Mesenchymal Stem Cells for Aortic Aneurysm

Introduction - We have previously reported effectiveness of autologous bone marrow derived mesenchymal stem cells (BM-MSCs) for regression of aortic aneurysm (AA). However, they have various disadvantage in terms of cell quality and quantity. Meanwhile, it is known that the stem cell has ability of immunological tolerance. The purpose of this study were to investigate whether treatment of allogeneic MSCs is effective for angiotensin II (AT II)-induced apolipoprotein E-deficient (apoE−/−) already-formed AA mouse. Methods - To identify characterization of each MSCs, the surface markers were checked by flow cytometry. AA was induced in apoE−/− mice by Ang II-infusion for 28 days through subcutaneously implanted osmotic mini-pump. The mice were randomly divided into 3 groups: (ⅰ) 0.2ml saline was intra-veinously injected (n=10, group S), (ⅱ) 1 × 106 autologous BM-MSCs(isolated from C57BL/6) in 0.2 ml saline (n=10, group Au), (ⅲ) 1 × 106 allogeneic BM-MSCs (isolated from BALB/C) (n=10, group Al) via tail vein. Mice were sacrificed at 2weeks after injection. Aortic diameter, matrix metalloproteinase (MMP)-2 and -9 enzymatic activity and cytokine concentrations were measured in obtained aortic tissue. Results - Flow cytometric analysis demonstrated that the both MSCs were similar in surface markers .The Aortic diameters of group Au and Al were significantly decreased compared to group S (group S vs Au vs Al; 2.29 vs 1.40 vs 1.36 (mm), p<0.01). The enzymatic activities of MMP-2 and -9 were significantly reduced in both MSCs groups (active-MMP2: group S vs Au vs Al; 0.45 vs 0.28 vs 0.24 (unit/ml), p<0.05 amd active-MMP9: group S vs Au vs Al; 0.34 vs 0.16 vs 0.14 (unit/ml), p<0.05). Inflammatory cytokines were down-regulated in the both MSC groups (interleukin-6: group S vs Au vs Al; 3399.5 vs 1475.6 vs 937.5 (pg/ml), p<0.01 and monocyte chemotactic protein-1: group S vs Au vs Al; 352.7 vs 208.0 vs 165.1 (pg/m), p<0.05).Insulin-like growth factor (IGF)-1 and tissue inhibitor of metalloproteinase (TIMP)-2 were up-regulated in the MSC groups (IGF-1: group S vs Au vs Al; 2.0 vs 4.7 vs 5.7 (pg/ml), p<0.01; TIMP-2: group S vs Au vs Al; 4.0 vs 9.5 vs 11.1 (pg/ml), p<0.01). Both MSCs injection inhibited infiltration of M1 macrophages in the aortic tissue. Conclusion - Our results suggest that the therapeutic effect of allogeneic MSCs might be similar to that of autologous MSCs for treatment of AA. Allogeneic MSCs injection could provide an alternative therapeutic strategy for the treatment of AA.

Mesenchymal Stem Cells Exhibit Both a Proinflammatory and Anti-Inflammatory Effect on Saccular Aneurysm Formation in a Rabbit Model.

Several studies have demonstrated a potential interaction between mesenchymal stem cells (MSCs) and saccular aneurysms. In this study, we sought to determine whether allogenic bone marrow-derived MSCs had the ability to prevent aneurysm formation in a known rabbit elastase aneurysm model. MSCs were injected intravenously in experimental rabbits at the time of surgical creation and two weeks postcreation and compared with control rabbits receiving vehicle injection. Angiography was used to compare aneurysm measurements four weeks postcreation, and aneurysms were harvested for histological properties. Serum was collected longitudinally to evaluate cytokine alterations. Serum from control animals was also utilized to perform in vitro tests with MSCs to compare the effect of the serologic environment in animals with and without aneurysms on MSC proliferation and cytokine production. While aneurysm morphometric comparisons revealed no differences, significant cytokine alterations were observed in vitro and in vivo, suggesting both anti-inflammatory and proinflammatory processes were occurring in the presence of MSCs. Histological analyses suggested that tunica intima hyperplasia was inhibited in the presence of MSCs.

BMSCs pre-treatment ameliorates inflammation-related tissue destruction in LPS-induced rat DIC model

This study aimed to investigate the effect of bone marrow-derived mesenchymal stem cells (BMSCs) on disseminated intravascular coagulation (DIC) model rats and to further explore the underlying mechanism. A rat model of lipopolysaccharide (LPS)-induced DIC was successfully established, as indicated by impaired plasma hemostatic parameters and damaged organ functions in rats. Importantly, pre-treatment with rat allogeneic BMSCs before LPS injection significantly alleviated systemic intravascular coagulation, reduced plasma levels of organ dysfunction indicators and pro-inflammatory cytokines, suppressed fibrin microthrombi formation, ameliorated liver, heart, and renal injuries, and increased 24-hour survival rates in LPS-induced DIC rats. The protection of BMSCs against DIC was in a moderately dose-dependent manner. Further investigation revealed that BMSCs co-cultured with peripheral blood mononuclear cells (PBMCs) significantly inhibited the LPS-stimulated PBMCs proliferation and the release of pro-inflammatory cytokines from PBMCs. Of note, upregulation of immunosuppressive factors including indoleamine 2,3-dioxygenase and interleukin-10, which was induced by interferon-γ, contributed to BMSCs-mediated inhibition of LPS-stimulated PBMCs proliferation. These effects do not depend on the direct cell–cell contact. In conclusion, BMSCs pre-treatment ameliorates inflammation-related tissue destruction in LPS-induced DIC model rats. The protection of BMSCs may be attributed to their anti-inflammatory and immunomodulatory properties, which render BMSCs a promising source for stem cell-based therapeutic approaches in inflammation-related DIC.

Intravenous Administration of Bone Marrow-Derived Mesenchymal Stem Cell, but not Adipose Tissue-Derived Stem Cell, Ameliorated the Neonatal Hypoxic-Ischemic Brain Injury by Changing Cerebral Inflammatory State in Rat.

Perinatal hypoxic-ischemic (HI) brain injury occurs in 1 in 1,000 live births and remains the main cause of neurological disability and death in term infants. Cytotherapy has recently emerged as a novel treatment for tissue injury. In particular, mesenchymal stem cells (MSCs) are thought to have therapeutic potential, but little is known about the differences according to their origin. In the current study, we investigated the therapeutic effects and safety of intravenous injection of allogeneic bone marrow-derived MSCs (BM-MSCs) and adipose-derived stem cells (ADSCs) in a rat model of HI brain injury. HI models were generated by ligating the left carotid artery of postnatal day 7 Wistar/ST rats and exposing them to 8% hypoxia for 60 min. Bone marrow and adipose tissue were harvested from adult green fluorescent protein transgenic Wistar rats, and cells were isolated and cultured to develop BM-MSCs and ADSCs. At passaging stages 2–3, 1 × 105 cells were intravenously injected into the external right jugular vein of the HI rats at 4 or 24 h after hypoxia. Brain damage was evaluated by counting the number of cells positive for active caspase-3 in the entire dentate gyrus. Microglial isotypes and serum cytokines/chemokines were also evaluated. Distribution of each cell type after intravenous injection was investigated pathologically and bio-optically by ex vivo imaging (IVIS®) with a fluorescent lipophilic tracer DiR. The mortality rate was higher in the ADSC group compared to the BM-MSC group, in pups injected with cells 4 h after hypoxia. The number of active caspase-3-positive cells significantly decreased in the BM-MSC group, and the percentage of M1 microglia (a proinflammatory isotype) was also lower in the BM-MSC vs control group in the penumbra of the cortex. Moreover, BM-MSC administration increased anti-inflammatory cytokine and growth factor levels, while ADSCs did not. Each injected cell type was mainly distributed in the lungs and liver, but ADSCs remained in the lungs longer. Pathologically, pulmonary embolisms and diffuse alveolar hemorrhages were seen in the ADSC group. These results indicated that injection of allogeneic BM-MSCs ameliorated neonatal HI brain injury, whereas ADSCs induced severe lung hemorrhage and higher mortality.

An allogenic therapeutic strategy for canine spinal cord injury using mesenchymal stem cells.

This study was conducted to characterize canine bone marrow-derived mesenchymal stem cells (BMSCs); in vivo tracking in mice, and therapeutic evaluation in canine clinical paraplegia cases. Canine BMSCs were isolated, cultured, and characterized in vitro as per International Society for Cellular Therapy criteria, and successfully differentiated to chondrogenic, osteogenic, and adipogenic lineages. To demonstrate the homing property, the pGL4.51 vector that contained luciferase reporter gene was used to transfect BMSCs. Successfully transfected cells were injected around the skin wound in mice and in vivo imaging was done at 6, 12 and 24 hr post MSCs delivery. In vivo imaging revealed that transfected BMSCs migrated and concentrated predominantly toward the center of the wound. BMSCs were further evaluated for allogenic therapeutic potential in 44 clinical cases of spinal cord injuries (SCI) and compared with conventional therapy (control). Therapeutic potential as evaluated by different body reflexes and recovery score depicted significantly better results in stem cell-treated group compared to control group. In conclusion, allogenic canine BMSCs can serve as potent therapeutic candidate in cell-based therapies, especially for diseases like SCI, where the conventional medication is not so promising.

Retrospective analysis of local injection site adverse reactions associated with 230 allogenic administrations of bone marrow-derived mesenchymal stem cells in 164 horses.

Bone marrow-derived mesenchymal stem cells (BM-MSCs) are frequently used in the treatment of musculoskeletal injuries. Fully characterised cells that are readily available for use is optimum. Allogenic BM-MSCs can satisfy the need for rapid treatment, however, their safety has been questioned. Objectives were to characterise BM-MSCs from an adult donor horse, in vitro, and to identify and describe adverse reactions that occurred following their injection into other horses. We hypothesised that BM-MSCs capable of proliferation, differentiation and lacking MHC II from one donor could be implanted into another individual without significant adverse reactions and the frequency of adverse reactions in clinical cases would be similar to that previously reported for autologous BM-MSCs. Retrospective clinical study. BM-MSCs were proliferated and characterised from one donor and cryopreserved for clinical use. Medical records for horses injected with allogenic BM-MSCs from this donor at a single hospital were used. After routine lameness exam, lesions were identified using diagnostic ultrasound or MRI. Post injection reaction was defined as increased pain, swelling, or heat at or near injection site, or increased lameness. Treatments required for each reaction were noted. BM-MSCs proliferated and underwent differentiation. Cells were found to be negative for MHC-II (<2%) and were viable after cryopreservation and shipping. Ten of 230 (4.35%) injections were noted to be associated with an adverse reaction. Adverse reactions occurred in synovial structures (n = 3) and in soft tissues (n = 7). This investigation could underestimate the number and severity of reactions. Mild reactions, such as synovitis, may have been missed. Also, anti-inflammatory drugs could overshadow mild reactions, making them less likely to be detected. Fully characterised allogenic BM-MSCs originating from a single donor horse can be administered to horses with soft tissue injuries with a low rate of adverse reaction. The Summary is available in Portuguese - see Supporting Information.

Potential Effect of Bone Marrow-Derived Mesenchymal Stem Cells on Zymogen Granules and Rab3D in Parotid Salivary Glands of Senile Albino Rats

Background: Age-related changes of salivary glands affect their functions and structure. Rab3D is localized to zymogen granules (ZGs) and plays a critical role in secretory granules maturation and exocytosis. Nowadays, application of life sciences is used in tissue regeneration. Objective: The current study was designed to explore the possible effect of bone marrow-derived mesenchymal stem cells (BM-MSCs) on the function and morphology of ZGs in parotid glands of senile rats.Design: Fourteen clinically healthy male albino rats were used. Seven adult rats aged 6-8months were named as Control adult (Group I). The remaining seven rats aged 18-20 months (old age). For each old age rat the parotid gland of the left side represented the Old age untreated (Group II) and injected by 0.2 ml phosphate buffered saline (PBS). While, the parotid gland of the right side represented the Old age treated by stem cells (Group III) and injected by 1-1.5 x 106 BM-MSCs in 0.2 ml PBS. At the end of the experimental period (after four weeks), all the rats were euthanized and the parotid salivary glands were dissected out. The sections were prepared for histological, immunohistochemical and ultra-structural examinations in addition to real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) analysis. Statistical analysis: Data obtained from histomorphometric analysis and qRT-PCR assay were statistically described in terms of mean ± standard deviation (± SD).Results: Histological examination of Group I showed normal histological features of the terminal secretory units. Group II revealed that most of the acini were distorted and showed fatty degeneration. The oncocytes were noticeable. Group III illustrated more or less normal histological features. Immuno-expression of Rab3D was strong in Group I and Group III but mild in Group II. The ultra-structural examination of Group I showed apparently large and spherical ZGs. Group II revealed apparently small and ellipsoid ZGs. Lamellated lipofuscin granules were observed. Group III demonstrated obviously large and well defined spherical ZGs. The statistical results revealed that the highest means of Rab3D immunoreactivity area % and relative Rab3D gene expression level were demonstrated in Group I, followed by Group III, and the least values were for Group II. Conclusions: Allogeneic BM-MSCs have positive effect on age-related changes of parotid salivary glands by restoring the terminal secretory units’ structure, Rab3D expression as well as ZGs structure and function.

Bone Marrow-Derived Mesenchymal Stem Cells-Mediated Protection Against Organ Dysfunction in Disseminated Intravascular Coagulation Is Associated With Peripheral Immune Responses.

Disseminated intravascular coagulation (DIC) is a fatal thrombohemorrhagic disorder. Bone marrow-derived mesenchymal stem cells (BMSCs) are multipotent stem cells that have tremendous therapeutic effect. Our aim was to explore whether the immune mechanisms were associated with BMSCs-afforded protection against DIC. We generated a rat model of DIC by lipopolysaccharide (LPS, 3 mg/kg) injection via the tail vein. In the treatment group, rats were pre-treated with 1 × l03 , 1 × l04 , 1 × l05 , and 1 × l06 allogeneic BMSCs before LPS injection. Blood sample was withdrawn from the abdominal aorta at 0 (before), 4, and 8 h after LPS injection and used for biochemical analyses. After experiments, the mice were sacrificed and their organs were harvested and observed by H&E and PTAH staining. Continuous infusion of LPS into the rats gradually impaired the hemostatic parameters and damaged organ functions. However, pre-treatment with BMSCs dose-dependently improved the hemostatic parameters. Meanwhile, the treatment significantly suppressed the fibrin microthrombi formation and alleviated liver, heart, lung, and renal injuries. Flow cytometry analysis demonstrated that BMSCs pre-treatment inhibited LPS-induced upregulation of CD3+ CD8+ T cells and CD3+ /CD161a+ NKT cells in the peripheral blood. BMSCs pre-treatment reversed the upregualtion of the B-cell population and the percentage of CD43+ /CD172a+ monocytes in the DIC models. Finally, BMSCs pre-treatment decreased the levels of tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), interleukin-1β (IL-1β), and interleukin-6 (IL-6) and increased the levels of interleukin-10 (IL-10) in LPS-induced DIC models. Pre-treatment with BMSCs can reduce coagulation and alleviate organ dysfunction via peripheral immune responses in LPS-induced DIC rat model. J. Cell. Biochem. 118: 3184-3192, 2017. © 2017 Wiley Periodicals, Inc.

Effect of Bone Marrow Versus Adipose Tissue Derived Mesenchymal Stem Cells on the Pancreas of Streptozotocin-Induced Diabetes Mellitus Type I in Adult Male Rats (Histological Study)

Background: Diabetes Mellitus (D.M.) is a major health problem affecting more than 200 million worldwide. The ideal treatment for autoimmune type I diabetes is regeneration of endogenous β-cells which could be achieved by mesenchymal stem cells transplantation.Aim of the work: This work aimed to compare the effect of intravenous bone marrow derived mesenchymal stem cells (BMSCs) and adipose tissue derived mesenchymal stem cells (AMSCs) on Streptozotocin (STZ)-induced type I diabetes in albino rats. Material and Methods: Fifty albino male rats were divided into 4 groups; control, diabetic, BMSCs treated and AMSCs treated. Treated groups were intravenously given 1 ml PKH26 labeled allogenic BMSCs or AMSCs suspended in phosphate buffered saline, respectively. Animals of all groups were sacrificed 2 weeks after stem cells administration. Sections from control and treated groups were examined by fluorescence microscope. Sections from all groups were immunohistochemically stained to detect insulin and proliferating cell nuclear antigen (PCNA). Mean area percent of insulin and number of PCNA positive reactions were measured and statistically analyzed.Results: Diabetic rats showed cell death and congested blood vessels in both exocrine and endocrine pancreas. Treated groups revealed homing of stem cells in pancreas after their transplantation. Moreover, nearly normal histological features were seen in AMSCs treated group. Studying the treated groups immunohistochemically, revealed increase in insulin and PCNA positive reactions when compared to diabetic group with more increase in AMSCs treated group than BMSCs treated group. Conclusion: Intravenous AMSCs could be more effective than BMSCs in treatment of STZ-induced type I diabetes.