Electrical stimulation (ES) is known to guide the development and regeneration of many tissues. Use of low-frequency ES for therapeutic purposes has been increasing during the last decades. Mesenchymal stem cells (MSCs) represent an appealing alternative cell source for cartilage repair. There are studies that induce differentiation into cartilage cells by treating the growth factors in stem cells or altering the properties of stem cells by genetic modification. In this study, we exposed equine adipose tissue-derived MSCs (eAD-MSCs) to ES and assessed changes in the chondrogenic differentiation potential. The cells obtained from equine adipose tissue attached to culture plates and expanded in vitro. Flow cytometric analysis at third passage indicated that the cells were strongly positive for CD44, CD90, and CD105, but negative for CD13, CD34, and CD45. Next, ES was applied to eAD-MSCs cultured under condition of high-density micromass under ES of 10 V/cm, with duration of 10 ms and a frequency of 2.0 Hz for three days. Gene expression of chondrogenic markers such as collagen type II, Aggrecan, and Sox9 was analyzed at three days of ES. As a result, we observed the differentiation potential of eAD-MSCs into chondrocytes by specific ES in absence of exogenous growth factors. We also found that ES upregulated the expression of heat shock protein 70, which affects cartilage formation. This study may contribute to the differentiation of MSCs into chondrogenic lineage under specific ES condition.

Mesenchymal stem cells (MSCs) are capable of differentiating into mesenchymal tissue such as bone, cartilage, muscle, and adipose, and have been isolated and characterized from various species. Deer adipose tissue-derived MSCs (dAD-MSCs) have not been studied and deer bone marrow-derived MSCs (dBM-MSCs) have not been fully characterized. In this study, we firstly isolated MSCs from deer tissues and then compared characteristics of dAD-MSCs and dBM-MSCs. dAD-MSCs and dBM-MSCs exhibited significant increase in proliferation under low-glucose DMEM culture condition during 20 and 10 passages consecutive passages, respectively. Both cells expressed cell surface markers such as CD73, CD90, and CD105, but did not express CD34 and CD45. Two types of cells expressed stemness markers (Oct4, Sox2, and Nanog) and exhibited differentiation potential into mesodermal lineages. Both cells exhibited osteogenic and chondrogenic differentiation potential, but poor adipogenic differentiation potential. Specifically, dAD-MSCs have a greater capacity for chondrogenic differentiation potential compared to dBM-MSCs. Collectively, we successfully isolated dAD-MSCs from deer for the first time. This study suggests that adipose tissue of deer could be used as a source of deer MSCs.

Mesenchymal stem cells (MSCs) are multipotent cells capable of replicating as undifferentiated cells, and have the potential of differentiating into mesodermal lineages. Goats are commonly used as animal models for bone tissue engineering to test the potential of stem cells for bone regeneration. Goat MSCs isolated from bone marrow (BM) or adipose tissue (AD) should be evaluated using in vitro assays, prior to their application in a tissue engineering project. In this study, we compared the stem cell properties of MSCs derived from goat AD, BM and ear skin tissue (ESK). As results, BM and ESK-MSCs exhibited a spindle-shaped morphology comparable to that of AD-MSCs. Especially, BM-MSCs could be cultured for significantly longer periods and exhibited the greatest expansion capacity, whereas AD-MSCs had the shortest culture time and lowest growth rate. Also, we compared differentiation potentials of AD, BM and ESK-MSCs into adipogenic, chondrogenic, and osteogenic lineages through specific staining and quantitative real-time RT-PCR. Collectively, we successfully isolated ESK-MSCs from goat for the first time. This study suggests that adult skin tissue of goat could be used as a source of goat MSCs. Further studies are needed to show the more information for establishment and fully characterization of goat ESK-MSCs.

Musculoskeletal disorders including fracture, tendonitis, osteoarthritis, and laminitis are common diseases in racehorses that can cause large economic losses in the racehorse industry. Mesenchymal stem cells (MSCs) are being applied as new clinical tools for treatment of musculoskeletal disorders of racehorses. To investigate the immunomodulatory effects of stem cell therapy, we analyzed the anti- and pro-inflammatory factors in peripheral blood mononuclear cells of racehorses before and after stem cell application using quantitative real-time RT-PCR. The expression levels of pro-inflammatory factors (CCL5, IFN-γ, IL-2, and IL-18) were decreased while those of anti-inflammatory factors (TIMP-1, IL-10, TGF-β1, and VEGF) were increased significantly after application of equine adipose tissue-derived MSCs (eAD-MSCs) to racehorses with fracture. Moreover, the expression levels of pro-inflammatory factors (IL-2, IL-18, and TNF-α) were decreased while those of anti-inflammatory factors (TIMP-1, TIMP-2, IL-10, TGF-β1, and VEGF) increased significantly after stem cell application of eAD-MSCs in racehorses with tendonitis. After evaluating immunomodulatory effects of stem cell therapy on equine musculoskeletal disorders such as fracture and tendonitis, our results showed that expression levels of pro-inflammatory factors were decreased, while those of anti-inflammatory factors increased significantly after stem cell application of eAD-MSCs. These findings suggest that the healing effects of the stem cell therapy might be due to its modulation of inflammatory factors.

Mesenchymal stem cells (MSCs), which are present in all tissues, can differentiate into cells with various specific functions. Recently, cell-based therapies using MSCs have been increasing in the veterinary research and related fields. In this study, we investigated the cellular morphology, proliferating capacities, expression of cell surface markers such as CD13, CD34, CD44, CD45, CD90, and CD105, mesodermal differentiation potentials, and expression of senescence-related markers of p53, p21, and telomerase reverse transcriptase in equine adipose tissue-derived MSCs (eAD-MSCs) after cryopreservation. The eAD-MSCs were analyzed immediately and after being frozen in liquid nitrogen for 1 year (< 1 year, G1) and more than 3 years (> 3 years, G2), respectively. After cryopreservation for 1 - 3 years, G2 eAD-MSCs showed similar cellular morphology, proliferating capacities, and expression of cell surface markers when compared with G1 eAD-MSCs. Moreover, cryopreservation did not affect the adipogenic, chondrogenic, or osteogenic differentiation potentials of G1 and G2 eAD-MSCs. Collectively, cryopreservation for (or over) 3 years maintained the stem cell phenotype and differentiation potentials of eAD-MSCs. These results will be an advantage that can be effectively used for future development of cell-based therapies.

Induced pluripotent stem cells (iPSCs) can be generated from adult cells. Somatic cells can be reprogrammed to form iPSCs by overexpressing transcription factors such as Oct4, Sox2, cMyc, and Klf4. To maintain undifferentiated state of iPSCs in vitro, cells have traditionally been maintained on mouse embryonic fibroblast feeders and passaged by enzymatic or mechanical dissociation methods. In this study, we compared the morphology and pluripotency of porcine iPSCs (piPSCs) after subsequent passaging using enzymatic and mechanical dissociation methods. Enzymatically and mechanically passaged piPSCs showed embryonic stem cell-like morphologies with compact cell adhesion and clear colony borders. In addition, alkaline phosphatase staining was positive for both enzymatically and mechanically passaged piPSCs. However, visual observation revealed that some colonies of enzymatically passaged piPSCs were spontaneously differentiated more than those of piPSCs mechanically passaged from 5 passage. Quantitative real-time RT-PCR demonstrated that enzymatically and mechanically passaged piPSCs expressed pluripotent genes such as Oct4, Sox2 and Nanog well at early passage. Immunofluorescent staining also confirmed that pluripotent markers such as Oct4, Sox2, and Nanog were positively expressed at early passage. However, expression levels of pluripotent genes in mechanically passaged piPSCs were also higher than those in enzymatically passaged piPSCs at early passage. Collectively, we found that mechanical passage method was better than enzymatic passage in terms of morphology and pluripotency of piPSCs at early passage. Further studies are needed to compare these dissociation methods with those obtained after more passages of piPSCs.

Mesenchymal stem cells (MSCs) have restricted life spans in vitro and can therefore only be expanded for a limited number of cell divisions before entering a senescent state and unequivocally stopping proliferation. Several types of cell culture systems have been used for large-scale expansion of MSCs. A recent trend in cell culture has been the change from serum-use to serum-supplement media. This study was conducted to investigate the proliferative effects of vegetable resources (VR) on equine adipose tissue-derived mesenchymal stem cells (eAD-MSCs) in the absence of serum and their possible mechanisms of action. Regulation of cell cycling is a key process involved in the fate of stem cells, including renewal and differentiation. In this study, we observed that the viability of eAD-MSCs was increased significantly when treated with VR under serum-free conditions. We also observed that expression levels of cell cycling-related proteins such as p53 and p21 were decreased, and proliferating cell nuclear antigen increased significantly in response to treatment with VR in eAD-MSCs under serum-free conditions. Furthermore, expression levels of cell survival-related proteins were increased in response to treatment with VR in eAD-MSCs under serum-free conditions. Therefore, our results suggest that VR promotes proliferation of eAD-MSCs under serum-free conditions.

Mesenchymal stem cells (MSCs) are multipotent cells able to differentiate into several cell lineages, which has implications for cell therapy and reproductive biotechnologies. Although MSCs have been isolated from many species, including humans and animals, there is limited data on MSCs from large ruminants, such as bovines. In this study, we tried to isolate and characterize bovine tongue tissue-derived MSCs (boT-MSCs) by investigating phenotype morphology, performing proliferation properties, and determining cell surface marker expression patterns, self-renewal, and differentiation potentials. As a result, the boT-MSCs were successfully isolated by collagenase digestion and maintained proliferative capacity until 20 passages. Moreover, the boT-MSCs expressed pluripotency markers (OCT3/4, SOX2, and NANOG) and MSC-specific surface markers including CD44, CD90, and CD105, but not CD45 and MHC-II. The boT-MSCs could also differentiate into mesodermal (adipocyte, osteocyte, and chondrocyte) cell lineages. Our results suggest that the tongues of bovines could be used as a source of MSCs.

본 연구는 칼슘과 철 그리고 철과 아연의 상호작용과 신뢰성있는 영양정보를 제공하기 위해 영양강조표시제품중 칼슘, 철, 아연의 함량을 분석하고 표시량과 분석값을 비교하였다. 칼슘, 철, 아연을 강조표시한 제품(시리얼, 과자, 두유, 초콜릿가공품, 기타코코아가공품, 당류가공품, 과줆채음료, 고형차) 총 42건을 수거하였으며 칼슘, 철, 아연은 무기성분의 건식분해법으로 전 처리한 후 Inductively Coupled Plasma Spectrometer (ICP)로 실험하였다. 칼슘이 강조표시된 제품 42건에 대한 표시량 비율은 87~176%이 었으며, 철이 강조표시된 제품 13건에 대한 표시량 비율은 84~167%, 아연이 강조표시된 제품 6건에 대한 표시량비율은 98~275%였다. 모든 분석값이 표시량 대비 80% 이상으로 식품 등의 표시기준을 충족하였다. 칼슘과 철의 상호작용은 한국인의 칼슘 섭취가 1일 권장량의 68.7% 수준으로 부족하므로 칼슘의 과다섭취로 인한 철의 흡수방해는 우려할 만한 상황은 아닌 것으로 보인다. 또한, 분석한 영양강조표시제품의 철과 아연의 함량 비율이 1.53:1이 최대였으므로 철의 과다섭취로 인한 아연의 흡수방해는 우려할 만한 상황은 아니었다. 그러나 업체에서는 칼슘, 철, 아연을 강조한제품을 생산할 시에 영양소의 상호작용을 고려하여 생산을 해야 할 것이고 소비자에게 정확한 영양정보를 제공하고 적정량의 영양섭취를 위해서는 지속적인 모니터링을 통해 식품표시에 대한 주기적인 관리가 필요할 것으로 생각된다.

In this study, to improve the in vitro development of various cells including cloned embryos, the effects that isoproterenol and melatonin have on in vitro development of porcine parthenogenetic oocytes were investigated. Parthenogenetic activation was induced with electrical stimulation, BSA and 6-DMAP treatment. 10-7 M of melatonin and isoproterenol (10-10, 10-12 and 10-14 M) were supplemented for in vitro maturation (IVM) and in vitro culture (IVC) medium, with different concentrations. When isoproterenol and melatonin were supplemented in IVM medium with different concentrations, there was no significant (P<0.05) difference of maturation rate in the treatment groups as well as in that of only melatonin. As isoproterenol and melatonin were supplemented in IVM medium with different concentrations, blastocyst rates of isoproterenol 10-12 M treatment group (37.1%) were significantly (P<0.05) higher than control group (26.0%). Isoproterenol and melatonin were supplemented in IVC medium with different concentrations, then the cleavage rate of 10-12 M isoproterenol treatment group (82.2%) was significantly (P<0.05) higher than the group that melatonin was only supplemented (70.9%). There was no difference of blastocyst rate between the treatment groups. When isoproterenol and melatonin were supplemented for IVM+IVC medium with different concentrations, the cleavage rate of 10-12 M isoproterenol treatment group (92.5%) was significantly (P<0.05) higher than the control group (82.8%) and the group that melatonin was only treated (81.6%). The blastocyst rate of 10-12 M as 45.6% was significantly (P<0.05) higher than control group (25.2%) and melatonin treatment group (31.2%). The cell number of blastocyst in 10-12 M isoproterenol treatment group 35.5±3.4 was significantly (P<0.05) highest. The results of this study showed that the development rate of IVC when both isoproterenol and melatonin were supplemented was higher than when melatonin was only supplemented. Therefore, it is concluded that isoproterenol is rather effective in the activation of melatonin. 10-7 M melatonin and 10-12 M isoproterenol were considered suitable concentration.