Anti-cancer effects of DHP107 on canine mammary gland cancer examined through in-vitro and in-vivo mouse xenograft models.

BACKGROUND: Canine mammary gland cancer (CMGC) is a common neoplasm in intact bitches. However, the benefit of adjuvant chemotherapy is unclear. The aim of this study was to investigate the anti-proliferative effects of paclitaxel on CMGC in in-vitro and in-vivo settings. RESULTS: Paclitaxel dose-dependently inhibited viability and induced G2/M phase cell cycle arrest and apoptosis in both primary and metastatic CMGC cell lines (CIPp and CIPm). In animal experiments, the average tumour volume decreased significantly in proportion to the administered oral paclitaxel dose. By examining tumour tissue using a TUNEL assay and immunohistochemical staining with anti-CD31 as a marker of endothelial differentiation, respectively, it was confirmed that oral paclitaxel induced apoptosis and exerted an anti-angiogenetic effect in tumour tissues. Further, downregulation of cyclin D1 in tumour tissues suggested that oral paclitaxel induced cell cycle arrest in tumour tissues in-vivo. CONCLUSIONS: Our results suggest that paclitaxel may have anti-cancer effects on CMGC through cell cycle arrest, induction of apoptosis, and anti-angiogenesis. This study could provide a novel approach to treat CMGC.

Effects of high fat diet-induced obesity on pathophysiology, immune cells, and therapeutic efficacy in systemic lupus erythematosus.

Prior studies have suggested a strong link between obesity and autoimmune diseases. This study aimed to evaluate the effects of high fat diet (HFD)-induced obesity on the disease pathogenesis, immune cell infiltration, and therapeutic efficacy in systemic lupus erythematosus (SLE). Treatment with methylprednisolone significantly increased the survival in the control diet group, but not in the HFD group. An HFD significantly increased the incidence of severe proteinuria and glucose intolerance. Regardless of the diet, treatment with methylprednisolone significantly decreased the serum levels of anti-dsDNA antibodies, IL-2, IL-10, and interferon γ-induced protein 10 (IP-10), and improved the renal pathology scores. Treatment with methylprednisolone significantly lowered the serum levels of IL-6, MCP-1, and TNF-α in the control diet group, but not in the HFD group. HFD significantly increased the proportions of CD45+ and M1 cells and significantly decreased the proportion of M2 cells in white adipose tissue; methylprednisolone treatment significantly rescued this effect. In the HFD group, methylprednisolone treatment significantly decreased the M1:M2 and increased the Foxp3+:RORγt+ cell in the spleen compared with the untreated group. These data improve our understanding of the effect of HFD on the therapeutic efficacy of corticosteroids in SLE treatment, which could have clinical implications.

Withaferin A mitigates metastatic traits in human oral squamous cell carcinoma caused by aberrant claudin-1 expression.

Abnormal expression of claudin-1 (CLDN1) has important roles in carcinogenesis and metastasis in various cancers. The role of CLDN1 in human oral squamous cell carcinoma (OSCC) remains unknown. Here, we report the functional role of CLDN1 in metastasis of human OSCC, as a potential target regulated by withaferin A. From gene expression profiling with microarray technology, we found that the majority of notable differentially expressed genes were classified into migration/invasion category. Withaferin A impaired the motility of human OSCC cells in vitro and suppressed metastatic nodule formation in an in vivo metastasis model, both associated with reduced CLDN1. CLDN1 overexpression enhanced metastatic nodule formation in vivo, resulting in severe metastatic lesions in lung tissue. Moreover, CLDN1 expression was positively correlated to lymphatic metastasis in OSCC patients. The impaired motility of human OSCC cells upon withaferin A treatment was restored by CLDN1 overexpression. Furthermore, upregulation of let-7a induced by withaferin A was inversely correlated to CLDN1 expression. Overall, these give us an insight into the function of CLDN1 for prognosis and treatment of human OSCC, substantiating further investigation into the use of withaferin A as good anti-metastatic drug candidate.

Preparation of Stretchable Nanofibrous Sheets with Sacrificial Coaxial Electrospinning for Treatment of Traumatic Muscle Injury.

Traumatic muscle injury with massive loss of muscle volume requires intramuscular implantation of proper scaffolds for fast and successful recovery. Although many artificial scaffolds effectively accelerate formation and maturation of myotubes, limited studies are showing the therapeutic effect of artificial scaffolds in animal models with massive muscle injury. In this study, improved myotube differentiation is approved on stepwise stretched gelatin nanofibers and applied to damaged muscle recovery in an animal model. The gelatin nanofibers are fabricated by a two-step process composed of co-axial electrospinning of poly(ɛ-caprolactone) and gelatin and subsequent removal of the outer shells. When stepwise stretching is applied to the myoblasts on gelatin nanofibers for five days, enhanced myotube formation and polarized elongation are observed. Animal models with volumetric loss at quadriceps femoris muscles (>50%) are transplanted with the myotubes cultivated on thin and flexible gelatin nanofiber. Treated animals more efficiently recover exercising functions of the leg when myotubes and the gelatin nanofiber are co-implanted at the injury sites. This result suggests that mechanically stimulated myotubes on gelatin nanofiber is therapeutically feasible for the robust recovery of volumetric muscle loss.

Dual delivery of stem cells and insulin-like growth factor-1 in coacervate-embedded composite hydrogels for enhanced cartilage regeneration in osteochondral defects.

Exogenous dual delivery of progenitor cell population and therapeutic growth factors (GFs) is one of alternative tissue engineering strategies for osteochondral tissue regeneration. In the present study, an implantable dual delivery platform was developed using coacervates (Coa) (i.e., a tertiary complex of poly(ethylene argininylaspartate diglyceride) (PEAD) polycation, heparin, and cargo insulin-like growth factor-1 (IGF-1), in thiolated gelatin (gelatin-SH)/ poly(ethylene glycol) diacrylate (PEGDA) interpenetrating network (IPN) hydrogels. Since Coa is able to protect cargo GF and maintain its long-term bioactivity, it is speculated that Coa-mediated delivery of chondrogenic factor IGF-1 with the aid of adipose-derived stem cells (ADSCs) would synergistically facilitate osteochondral tissue repair during physiological regeneration process. Our results indicate that gelatin-SH/PEGDA IPN hydrogels demonstrated biocompatibility and mechanical properties for a possible long-term transplantation, and PEAD-base Coa exhibited a sustained release of bioactive IGF-1 over 3 weeks. Subsequently, released IGF-1 from Coa could effectively induce chondrogenic differentiation of embedded ADSCs in the hydrogel, by showing enhanced glycosaminoglycan deposition and expression of chondrogenesis-associated genes. More importantly, at 12 weeks post-implantation in a rabbit full thickness osteochondral defect model, the quality of regenerative tissues in both chondral and subchondral layers was significantly improved in dual delivery of ADSC and IGF-1 in Coa encapsulated in gelatin-SH/PEGDA IPN hydrogels, as compared with a single delivery of ADSC only and a dual delivery without Coa. Therefore, we conclude that our Coa-embedded composite hydrogel platform could effectively augment osteochondral tissue regeneration holds promise for a feasible osteoarthritis therapeutic application.

Oridonin induces the apoptosis of mucoepidermoid carcinoma cell lines in a myeloid cell leukemia­1­dependent manner.

Oridonin, an active diterpenoid isolated from Rabdosia rubescens, has been reported to exhibit anticancer activities in several tumors. The aim of the present study was to investigate the anticancer effects and molecular mechanisms of oridonin in mucoepidermoid carcinoma (MEC). Treatment with oridonin induced the apoptosis of MC­3 and YD­15 cell and inhibited the expression of myeloid cell leukemia­1 (MCL­1) through the regulation of the protein level through post­translational regulation in these cell lines. Oridonin significantly increased the expression level of truncated Bid (t­Bid) as a downstream target of MCL­1 and subsequently decreased the mitochondrial membrane potential. The ectopic expression of MCL­1 protein was sufficient to reverse the induction of apoptosis and the increased t­Bid expression induced by oridonin in both cell lines. Taken together, these results suggest that oridonin exerts an apoptotic effect through the modulation of MCL­1 and t­Bid in human MEC cell lines and may thus be a potential anticancer drug candidate for the treatment of human MEC.

TSG-6 in extracellular vesicles from canine mesenchymal stem/stromal is a major factor in relieving DSS-induced colitis.

Adipose tissue derived mesenchymal stem/stromal cell (ASC)-derived extracellular vesicles (EV) have been reported to be beneficial against dextran sulfate sodium (DSS)-induced colitis in mice. However, the underlying mechanisms have not been fully elucidated. We hypothesize that the tumor necrosis factor-α-stimulated gene/protein 6 (TSG-6) in EVs is a key factor influencing the alleviation of colitis symptoms. DSS-induced colitis mice (C57BL/6, male, Naïve = 6, Sham = 8, PBS = 8 EV = 8, CTL-EV = 8, TSG-6 depleted EV = 8) were intraperitoneally administered EVs (100 ug/mice) on day 1, 3, and 5; colon tissues were collected on day 10 for histopathological, RT-qPCR, western blot and immunofluorescence analyses. In mice injected with EV, inflammation was alleviated. Indeed, EVs regulated the levels of pro- and anti-inflammatory cytokines, such as TNF-α, IL-1ß, IFN-γ, IL-6, and IL-10 in inflamed colons. However, when injected with TSG-6 depleted EV, the degree of inflammatory relief was reduced. Furthermore, TSG-6 in EVs plays a key role in increasing regulatory T cells (Tregs) and polarizing macrophage from M1 to M2 in the colon. In conclusion, this study shows that TSG-6 in EVs is a major factor in the relief of DSS-induced colitis, by increasing the number of Tregs and macrophage polarization from M1 to M2 in the colon.

A placebo-controlled study comparing the efficacy of intra-articular injections of hyaluronic acid and a novel hyaluronic acid-platelet-rich plasma conjugate in a canine model of osteoarthritis.

BACKGROUND: The objective of this study was to assess the efficacy of intra-articular injections of hyaluronic acid (HA) and a novel, on-site conjugate of HA with autologous fibrinogen in platelet-rich plasma (HA-PRP) in a canine model of osteoarthritis (OA) METHODS: Twelve beagle dogs underwent a unilateral resection of the cranial cruciate ligament (CrCL) of the stifle joint. Clinical and radiographic signs of OA were confirmed in all dogs 8 weeks following CrCL resection and prior to treatment. The dogs were randomized into three groups: saline (n = 4), HA (n = 4), and HA-PRP (n = 4). Each dog received intra-articular injections of the respective substance into the affected joint at pre-determined time points. The dogs were assessed for adverse effects for 3 days after each injection and for lameness, pain, range of motion, kinetics, and radiographic OA severity prior to treatment and 3 months after injection. OA severity as determined by radiographic examination was not significantly different among the groups at any time point. The dogs were then humanely euthanatized and the stifle joint assessed by gross and histological examinations. RESULTS: Dogs treated with four weekly injections of HA or two biweekly injections of HA-PRP were significantly (p < 0.05) better than dogs treated with four weekly injections of saline at 2-, 4-, and 12-week time points based on a comfortable range of motion (CROM) and clinical lameness score. Gait analysis measuring symmetry and weight distribution on pressure sensor walkway showed significantly (p < 0.05) improved limb function for dogs treated with HA and HA-PRP compared with dogs treated with saline yet with better clinical outcome for the HA-PRP-treated group at 12 and 20 weeks follow-up. Gross and histological analysis of synovium and articular cartilage demonstrated significant (p < 0.05) improvement by both treatments groups compared to controls. There was however significantly (p < 0.05) less damage to the cartilage in the HA-PRP group compared to the HA-treated group. CONCLUSIONS: These data suggest that while injection of HA and HA-PRP may be sufficient for short-term amelioration of the symptoms associated with OA, treatment with HA-PRP conjugates may be superior, providing significantly better long-term cartilage preservation.

Contribution of p38 MAPK Pathway to Norcantharidin-Induced Programmed Cell Death in Human Oral Squamous Cell Carcinoma.

Norcantharidin (NCTD), a demethylated analog of cantharidin isolated from blister beetles, has been used as a promising anticancer agent; however, the underlying function of NCTD against human oral squamous cell carcinoma (OSCC) has not been fully understood. Here, this study was aimed to investigate the apoptotic effect and molecular targets of NCTD in human OSCC in vitro and in vivo. The anticancer effects of NCTD and its related molecular mechanisms were evaluated by trypan blue exclusion assay, live/dead assay, western blotting, 4-6-Diamidino-2-Phenylindole (DAPI) staining, flow cytometric analysis, Terminal Deoxynucleotidyl Transferase dUTP Nick end Labeling (TUNEL) assay, and immunohistochemistry. NCTD significantly inhibited cell growth and increased the number of dead cells in HSC-3 and HN22 cell lines. It induced the following apoptotic phenomena: (1) the cleavages of poly (ADP-ribose) polymerase and casepase-3; (2) increase in apoptotic morphological changes (nuclear condensation and fragmentation); (3) increase in annexin V-positive cells or sub-G1 population of cells. NCTD significantly activated the p38 mitogen-activated protein kinase (MAPK) pathway but inactivated the signal transducer and activator of transcription (STAT)3 pathway. A p38 MAPK inhibitor (SB203580) partially attenuated NCTD-induced programmed cell death (apoptosis) in both cell lines, whereas ectopic overexpression of STAT3 did not affect it. NCTD strongly suppressed tumor growth in the tumor xenograft bearing HSC-3 cells, and the number of TUNEL-positive cells increased in NCTD-treated tumor tissues. In addition, NCTD did not cause any histopathological changes in the liver nor the kidney. NCTD induced programmed cell death via the activation of p38 MAPK in OSCC. Therefore, these results suggest that NCTD could be a potential anticancer drug candidate for the treatment of OSCC.

Canine adipose tissue-derived mesenchymal stem cells pre-treated with TNF-alpha enhance immunomodulatory effects in inflammatory bowel disease in mice.

Canine inflammatory bowel disease (IBD) is an intractable autoimmune disorder that results in various gastrointestinal and systemic symptoms. Mesenchymal stem cells (MSCs), which release immunomodulatory factors such as tumor necrosis factor-α (TNF-α)-induced gene/protein 6 (TSG-6) and prostaglandin E2 (PGE2), have been suggested as an alternative therapeutic option for IBD treatment in veterinary medicine. Furthermore, although it is known that MSCs pre-treated with pro-inflammatory cytokines show enhanced anti-inflammatory properties via the secretion of soluble factors, the underlying mechanisms of IBD remain unclear. The aim of this study was to demonstrate the therapeutic effects and corresponding mechanisms of canine adipose tissue-derived (cAT)-MSCs stimulated with TNF-α in mouse models of IBD. Mice with dextran sulfate sodium (DSS)- or dinitrobenzene sulfonic acid (DNBS)-induced colitis were injected intraperitoneally with cAT-MSCs pre-treated with TNF-α. Colitis severity was assessed and colon tissues were collected for histopathological, enzyme-linked immunosorbent assay, and flow cytometry analysis. cAT-MSCs stimulated with TNF-α secreted higher concentrations of immunomodulatory factors such as TSG-6 and PGE2, which play a key role in inducing phenotypic alterations in macrophages. Consequently, TNF-α-pre-treated cAT-MSCs further regulated colonic inflammatory cytokines such as interleukin (IL)-1ß, IL-6, and IL-10, and ameliorated DSS- or DNBS-induced colitis in mice. Additionally, we demonstrated that M1 macrophages (F4/80+/iNOS+ cells) were decreased in colon tissues from mice treated with TNF-α-pre-treated cAT-MSCs, whereas M2 macrophages (F4/80+/CD206+ cells) were increased. These results may suggest a new cell-based therapeutic option for treating IBD.

Heme Oxygenase-1 is a Key Molecule Underlying Differential Response of TW-37-Induced Apoptosis in Human Mucoepidermoid Carcinoma Cells.

TW-37 is a small-molecule inhibitor of Bcl-2 family proteins, which can induce anti-cancer activities in various types of cancer. In the current study, we investigated the potential molecular mechanism underlying the differential response to TW-37-induced apoptosis in two human mucoepidermoid carcinoma (MEC) cell lines. The differential response and underlying molecular mechanism of human MEC cells to TW-37 was evaluated by trypan blue exclusion assay, western blotting, 4', 6-diamidino-2-phenylindole staining, annexin V/propidium iodide double staining, analysis of the sub-G1 population, human apoptosis array, and measurements of intracellular reactive oxygen species (ROS). TW-37 decreased cell viability and induced apoptosis in YD-15 cells, but not in MC3 cells. Proteome profiling using a human apoptosis array revealed four candidate proteins and of these, heme oxygenase-1 (HO-1) was mainly related to the differential response to TW-37 of YD-15 and MC3 cells. TW-37 also led to a significant increase in intracellular levels of ROS in YD-15 cells, which is associated with apoptosis induction. The ectopic expression of HO-1 recovered YD-15 cells from TW-37-induced apoptosis by reducing intracellular levels of ROS. The expression of HO-1 was reduced through both transcriptional and post-translational modification during TW-37-mediated apoptosis. We conclude that HO-1 is a potential indicator to estimate response to TW37-induced apoptosis in human MEC.

Inhibitory Effect of Olive Leaf Extract on Obesity in High-fat Diet-induced Mice.

BACKGROUND/AIM: The rapid increase in the number of people who are overweight or obese, which increases the risk of diseases and health problems, is becoming an important issue. Herein, we investigated whether olive leaf extract (OLE) has potent anti-obesity effects in high-fat induced mouse models. MATERIALS AND METHODS: C57BL/6 mice were randomized into normal control, high-fat diet (HFD), HFD with OLE, and HFD with garcinia groups and administered experimental diets for 12 weeks. Body weight and food intake were measured once per week and obesity-related biomarkers were evaluated in the serum and adipose tissue. RESULTS: OLE significantly suppressed weight gain, food efficiency ratio, visceral fat accumulation, and serum lipid composition in HFD-induced mice. Furthermore, the expression of adipogenesis- and thermogenesis-related molecules was decreased in the OLE-treated group. CONCLUSION: OLE prevents obesity development by regulating the expression of molecules involved in adipogenesis and thermogenesis.

Down-regulation and nuclear localization of survivin by sodium butyrate induces caspase-dependent apoptosis in human oral mucoepidermoid carcinoma.

OBJECTIVE: Sodium butyrate (NaBu) is a histone deacetylase inhibitor that possesses an apoptotic ability. However, the molecular mechanism by which NaBu induces apoptosis in human oral mucoepidermoid carcinoma (MEC), a type of salivary gland tumor, remains unclear. MATERIALS AND METHODS: The anticancer effects of NaBu and its related molecular mechanisms were determined by trypan blue exclusion assay, 4'-6-diamidino-2-phenylindole staining, live/dead assay, human apoptosis array, RT-PCR, western blotting, immunocytochemistry, preparation of nuclear fractions, and nude mice tumor xenograft. RESULTS: In this study, we found that NaBu inhibited growth and induced apoptosis in the human oral MEC cell lines MC3 and YD15 with acetylation of histone proteins H2A and H3. NaBu apparently down-regulated survivin protein, as evidenced by the results of the human apoptosis antibody array, and modulated it at the post-translational process. Interestingly, NaBu caused nuclear translocation of survivin protein in both cell lines. NaBu also resulted in decreased expression levels of Bcl-xL mRNA and protein, leading to induction of caspase-dependent apoptosis in human oral MEC cell lines. In addition, NaBu administration inhibited tumor growth in vivo at a dosage of 500 mg/kg/day, but it did not cause any hepatic or renal toxicity. CONCLUSION: This study provides new insights into the molecular mechanism of apoptotic actions by NaBu in human oral MEC and the basis of its clinical application for the treatment of human oral MEC.

Application of alginate microbeads as a carrier of bone morphogenetic protein-2 for bone regeneration.

Bone morphogenetic protein-2 (BMP-2) is commonly used to enhance bone regeneration. The potential of BMP-2 for bone regeneration varies according to the concentration and release kinetics on the implanted site. Therefore, it is important to determine appropriate carriers of BMP-2. However, no optimal delivery vehicles have been identified. In the present study, we used alginate microbeads as a delivery vehicle for BMP-2. Alginate microbeads can be implanted onto the disease site through surgery or injection. The objective of this study was to evaluate that the osteoinductive properties of BMP-2 are effective in alginate microbeads as a carrier. In this study, the release kinetics of BMP-2 in alginate microbeads was evaluated using an enzyme-linked immunosorbent assay. BMP-2 released from alginate microbeads induced high alkaline phosphatase activity in canine adipose tissue-derived mesenchymal stem cells. Injection of alginate microbeads with BMP-2 into mouse subcutaneous tissue, as well as surgical implantation into the 5-mm circular calvarial defects in rats, was conducted and the results showed extensive new bone formation. In conclusion, alginate microbeads can be utilized as an effective BMP-2 delivery vehicle for use in orthopedic surgery and as an injectable vehicle for a minimally invasive therapy. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 107B: 286-294, 2019.

Antitumor effect of TW-37, a BH3 mimetic in human oral cancer.

TW-37 is a small molecule B cell lymphoma-2 (Bcl-2) homology 3 mimetic with potential anticancer activities. However, the in vivo anti-cancer effect of TW-37 in human oral cancer has not been properly studied yet. Here, we attempted to confirm antitumor activity of TW37 in human oral cancer. TW-37 significantly inhibited cell proliferation and increased the number of dead cells in MC-3 and HSC-3 human oral cancer cell lines. TW-37 enhanced apoptosis of both cell lines evidenced by annexin V/propidium iodide double staining, sub-G1 population analysis and the detection of cleaved poly (ADP-ribose) polymerase and caspase-3. In addition, TW-37 markedly downregulated the expression of Bcl-2 protein, while not affecting Bcl-xL or myeloid cell leukemia-1. In vivo, TW-37 inhibited tumor growth in a nude mice xenograft model without any significant liver and kidney toxicities. Collectively, these data reveal that TW-37 may be a promising small molecule to inhibit human oral cancer.

Prostaglandin E2 secreted from feline adipose tissue-derived mesenchymal stem cells alleviate DSS-induced colitis by increasing regulatory T cells in mice.

BACKGROUND: Inflammatory bowel disease (IBD) is an intractable autoimmune disease, relatively common in cats, with chronic vomiting and diarrhea. Previous studies have reported that mesenchymal stem cells (MSCs) alleviate inflammation by modulating immune cells. However, there is a lack of research on cross-talk mechanism between feline adipose tissue-derived mesenchymal stem cells (fAT-MSCs) and immune cells in IBD model. Hence, this study aimed to evaluate the therapeutic effects of fAT-MSC on mice model of colitis and to clarify the therapeutic mechanism of fAT-MSCs. RESULTS: Intraperitoneal infusion of fAT-MSC ameliorated the clinical and histopathologic severity of colitis, including body weight loss, diarrhea, and inflammation in the colon of Dextran sulfate sodium (DSS)-treated mice (C57BL/6). Since regulatory T cells (Tregs) are pivotal in modulating immune responses and maintaining tolerance in colitis, the relation of Tregs with fAT-MSC-secreted factor was investigated in vitro. PGE2 secreted from fAT-MSC was demonstrated to induce elevation of FOXP3 mRNA expression and adjust inflammatory cytokines in Con A-induced feline peripheral blood mononuclear cells (PBMCs). Furthermore, in vivo, FOXP3+ cells of the fAT-MSC group were significantly increased in the inflamed colon, relative to that in the PBS group. CONCLUSION: Our results suggest that PGE2 secreted from fAT-MSC can reduce inflammation by increasing FOXP3+ Tregs in mice model of colitis. Consequently, these results propose the possibility of administration of fAT-MSC to cats with not only IBD but also other immune-mediated inflammatory diseases.

TSG-6 secreted by human adipose tissue-derived mesenchymal stem cells ameliorates severe acute pancreatitis via ER stress downregulation in mice.

BACKGROUND: Through recent studies, the onset of acute pancreatitis in pancreatic acinar cells (PACs) and the regulatory role of PACs in severe acute pancreatitis (SAP) have been revealed. During the early stages of pancreatitis, the endoplasmic reticulum (ER) in PACs undergoes significant changes, including swelling and vacuolization. In response to an increase in the extracellular stress in ER, PACs lose their functions, leading to cell apoptosis and inflammation response. The beneficial effects of human adipose tissue-derived mesenchymal stem cells (hAT-MSCs) on SAP have been well documented in previous studies. However, the underlying mechanism of their action remains controversial. METHODS: In this study, the therapeutic effects of intraperitoneally administered hAT-MSCs in a caerulein (50 µg/kg)- and lipopolysaccharide (LPS) (10 mg/kg)-co-induced SAP mouse model were evaluated. Inflammatory response and ER stress were measured in pancreatic tissue samples, and the beneficial effects were evaluated through quantitative reverse transcription polymerase chain reaction (qRT-PCR), western blot, and immunofluorescence analysis. RESULTS: Inflammatory response and ER stress were ameliorated following hAT-MSC injection, and the beneficial effects were observed in the absence of significant engraftment of hAT-MSCs. hAT-MSCs transfected with siRNA-targeting tumour necrosis factor-α-induced gene/protein 6 (TSG-6) were unable to inhibit ER stress and inflammation. In addition, TSG-6 from hAT-MSCs significantly suppressed ER stress-induced apoptosis and nuclear factor kappa B (NF-κB) activity in SAP model mice. CONCLUSIONS: TSG-6 secreted by hAT-MSCs protects PACs in SAP model mice via the inhibition of ER stress, as well as inflammatory responses. This study has revealed a new area for ER stress-targeted therapy in SAP patients.

Intra-Articular Injection of Alginate-Microencapsulated Adipose Tissue-Derived Mesenchymal Stem Cells for the Treatment of Osteoarthritis in Rabbits.

We investigated the effects of intra-articular injections of alginate-microencapsulated adipose tissue-derived mesenchymal stem cells (ASCs) during osteoarthritis (OA) development in a rabbit model of anterior cruciate ligament transection (ACLT). We induced OA in mature New Zealand white rabbits by bilateral ACLT. Stifle joints were categorised into four groups according to intra-articular injection materials. Alginate microbeads and microencapsulated ASCs were prepared using the vibrational nozzle technology. Two weeks after ACLT, the rabbits received three consecutive weekly intra-articular injections of 0.9% NaCl, alginate microbeads, ASCs, or microencapsulated ASCs, into each joint. Nine weeks after ACLT, we euthanised the rabbits and collected bilateral femoral condyles for macroscopic, histological, and immunohistochemical analyses. Macroscopic evaluation using the modified OA Research Society International (OARSI) score and total cartilage damage score showed that cartilage degradation on the femoral condyle was relatively low in the microencapsulated-ASC group. Histological analysis of the lateral femoral condyles indicated that microencapsulated ASCs had significant chondroprotective effects. Immunohistochemically, the expression of MMP-13 after the articular cartilage damage was relatively low in the microencapsulated-ASC-treated stifle joints. During the development of experimental OA, as compared to ASCs alone, intra-articular injection of microencapsulated ASCs significantly decreased the progression and extent of OA.

TSG-6 released from intraperitoneally injected canine adipose tissue-derived mesenchymal stem cells ameliorate inflammatory bowel disease by inducing M2 macrophage switch in mice.

BACKGROUND: Inflammatory bowel disease (IBD) is an intractable autoimmune disorder that markedly deteriorates one's quality of life. Mesenchymal stem cells (MSCs) alleviate inflammation by modulating inflammatory cytokines in inflamed tissues, and have been suggested as a promising alternative for IBD treatment in human and veterinary cases. Furthermore, tumor necrosis factor-α-induced gene/protein 6 (TSG-6) is a key factor influencing MSC immunomodulatory properties; however, the precise mechanism of TSG-6 release from canine MSCs in IBD remains unclear. This study aimed to assess the therapeutic effects of canine adipose tissue-derived (cAT)-MSC-produced TSG-6 in an IBD mouse model and to explore the mechanisms underlying the immunomodulatory properties. METHODS: Mice with dextran sulfate sodium-induced colitis were administered cAT-MSCs intraperitoneally; colon tissues were collected on day 10 for histopathological, quantitative real-time polymerase chain reaction, and immunofluorescence analyses. RESULTS: cAT-MSC-secreted TSG-6 ameliorated IBD and regulated colonic expression of pro- and anti-inflammatory cytokines such as tumor necrosis factor-α, interleukin-6, and interleukin-10. To investigate the effect of cAT-MSC-secreted TSG-6 on activated macrophages in vitro, a transwell coculture system was used; TSG-6 released by cAT-MSCs induced a macrophage phenotypic switch from M1 to M2. The cAT-MSC-secreted TSG-6 increased M2 macrophages in the inflamed colon in vivo. CONCLUSIONS: TSG-6 released from cAT-MSCs can alleviate dextran sulfate sodium-induced colitis by inducing a macrophage phenotypic switch to M2 in mice.

Injectable alginate-microencapsulated canine adipose tissue-derived mesenchymal stem cells for enhanced viable cell retention.

The purpose of this study was to establish an optimized protocol for the production of alginate-encapsulated canine adipose-derived mesenchymal stem cells (cASCs) and evaluate their suitability for clinical use, including viability, proliferation and in vivo cell retention. Alginate microbeads were formed by vibrational technology and the production of injectable microbeads was performed using various parameters with standard methodology. Microbead toxicity was tested in an animal model. Encapsulated cASCs were evaluated for viability and proliferation in vitro. HEK-293 cells, with or without microencapsulation, were injected into the subcutaneous tissue of mice and were tracked using in vivo bioluminescent imaging to evaluate the retention of transplanted cells. The optimized injectable microbeads were of uniform size and approximately 250 µm in diameter. There was no strong evidence of in vivo toxicity for the alginate beads. The cells remained viable after encapsulation, and there was evidence of in vitro proliferation within the microcapsules. In vivo bioluminescent imaging showed that alginate encapsulation improved the retention of transplanted cells and the encapsulated cells remained viable in vivo for 7 days. Encapsulation enhances the retention of viable cells in vivo and might represent a potential strategy to increase the therapeutic potency and efficacy of stem cells.