Tonsil-derived Mesenchymal Stem Cells Research Articles

Feasibility of autologous plasma gel for tonsil-derived stem cell therapeutics in hypoparathyroidism

Hypoparathyroidism is a deficiency of the parathyroid hormone (PTH) in the body. We previously reported the possibility of treating it using tonsil-derived mesenchymal stem cells (TMSCs) differentiated into PTH-releasing cells. The purpose of this study was to evaluate the feasibility of using autologous plasma gel as scaffold material in treatment of hypoparathyroidism with TMSC. We obtained plasma by venous sampling of autologous blood and centrifuged and fabricated the plasma gel using a sinusoidal pattern heating machine. After we created the hypoparathyroidism animal model, we administered undifferentiated TMSCs and TMSCs differentiated into parathyroid cells at each rat dorsum by intramuscular injection with and without the plasma gel. In the plasma gel groups, intact PTH was detected from on day 21 after TMSC injection; we did not detect intact PTH in the groups that were only transplanted with TMSCs during the entire experimental period. Serum calcium was higher and phosphorous was lower in the TMSC with plasma gel groups than in the groups with TMSCs alone. We detected PTH and chromogranin A in the TMSC-plasma gel-transplanted areas on immunohistochemistry and immunofluorescence stain. Plasma gel can be considered as a cell-delivery scaffold for treating hypoparathyroidism with tonsil-derived mesenchymal stem cells.

Double intratibial injection of human tonsil-derived mesenchymal stromal cells recovers postmenopausal osteoporotic bone mass

Background and aimsOsteoporosis, which is a disease characterized by weakening of the bone, affects a large portion of the senior population. The current therapeutic options for osteoporosis have side effects, and there is no effective treatment for severe osteoporosis. Thus, we urgently need new treatment strategies, such as topical therapies and/or safe and effective stem cell therapies. MethodsWe investigated the therapeutic potential of directly injecting human tonsil-derived mesenchymal stem cells (TMSC) into the right proximal tibias of ovariectomized postmenopausal osteoporosis model mice. Injections were given once (1×) or twice (2×) during the 3-month experimental period. At the end of the experiment, micro-computed tomographic images revealed some improvement in the proximal tibias and more significant improvement in the femoral heads of treated mice. ResultsOsteogenic effect was qualitatively and quantitatively more pronounced in TMSC/2×-treated mice. Furthermore, TMSC/2× mice exhibited significant recovery of the serum osteocalcin level, which is pathologically elevated in osteoporosis, and increased serum alkaline phosphatase, which indicates bone formation. TMSC therapy was generally well tolerated and caused no apparent toxicity in the experimental mice. Moreover, TMSC therapy reduced visceral fat. ConclusionOur results demonstrate that double injection of TMSC directly into the proximal tibia triggers recovery of osteoporosis, and thus could be a potential therapeutic approach for severe bone loss.

Tonsil-derived mesenchymal stem cell-embedded in situ crosslinkable gelatin hydrogel therapy recovers postmenopausal osteoporosis through bone regeneration.

We investigated therapeutic potential of human tonsil-derived mesenchymal stem cells (TMSC) subcutaneously delivered to ovariectomized (OVX) mice for developing more safe and effective therapy for osteoporosis. TMSC were isolated from tonsil tissues of children undergoing tonsillectomy, and TMSC-embedded in situ crosslinkable gelatin-hydroxyphenyl propionic acid hydrogel (TMSC-GHH) or TMSC alone were delivered subcutaneously to the dorsa of OVX mice. After 3 months, three-dimensionally reconstructed micro-computed tomographic images revealed better recovery of the femoral heads in OVX mice treated with TMSC-GHH. Serum osteocalcin and alkaline phosphatase were also recovered, indicating bone formation only in TMSC-GHH-treated mice, and absence in hypercalcemia or other severe macroscopic deformities showed biocompatibility of TMSC-GHH. Additionally, visceral fat reduction effects by TMSC-GHH further supported their therapeutic potential. TMSC provided therapeutic benefits toward osteoporosis only when embedded in GHH, and showed potential as a supplement or alternative to current therapies.

Optimization of Tenocyte Lineage-Related Factors from Mesenchymal Stem Cells

Category: Basic Sciences/Biologics Introduction/Purpose: Researchers should consider various potential factors that affect tenogenic differentiation of MSCs. Numerous experimental settings are associated with high cost and time. Response surface methodology (RSM), a component in the design of experiments (DOE), is gaining recognition as a powerful tool in optimizing conditions for the production of industrially important products such as chemicals and enzymes. The purpose of this study was to access the differential effects of transforming growth factor beta 3 (TGF-β3) on the tenogenesis of tonsil-derived mesenchymal stem cells (T-MSCs) and bone marrow-derived mesenchymal stem cells (BM-MSCs) using RSM. Methods: Bone marrow was collected from four patients (mean age: 79.0±2.2) and mononuclear cells were separated. The tonsillar tissues were collected from four patients (mean age: 7.6±0.6). After isolation of MSCs, they were treated with 5ng/ml and 10ng/ml of TGF-β3 with vehicle control. The full-factorial experimental design was employed to study the effect of tension based on T-MSCs. The design was composed of three levels being coded as -1, 0 and +1 and a total of 18 runs were carried out in duplicates to optimize the level of chosen variables, such as days and amount. A total of 84 trials were utilized and fitted with RSM; they were then used to obtain mathematical prediction models. Results: Exposure of TGF-β3 to T-MSCs and BM-MSCs resulted in an increase in the expression of SCX, TNMD, decorin, collagen I, and tenacin C. Most tenocyte lineage-related factors from T-MSCs and BM-MSCs presented a maximum increase in 2- 3 day treatment. Considering all of tenocyte lineage-related factors that we assessed, the predicted value of the factors was significantly induced at 2.7 ng/mL of TGF-β3 (p < 0.001) on 2.5-day culture (p = 0.001). (Fig A) Considering all of tenocyte lineage-related factors that we assessed, the predicted value of the factors was significantly induced on 2.3-day culture (p = 0.004) regardless of the concentration of TGF-β3. (Fig B) Conclusion: We demonstrated that tenocyte-like cells can be successfully differentiated from T-MSCs and BM-MSCs under TGF- β3 stimulation. This study demonstrated that T-MSCs and BM-MSCs in tenogenic stimulation with TGF-β3 have a similar tenogenic differentiation potential using RSM.

Tenogenic differentiation of mesenchymal stem cells

Category: Basic Sciences/Biologics Introduction/Purpose: Tendon repair has been a challenging issue for surgeons in treating. Although tissue engineering with mesenchymal stem cells (MSC) have been used for tendon repair in both in vivo and in vitro, the stem cells are obtained through invasive procedures, and there is usually a lack of adequate numbers for clinical use. The purpose of this study was to compare the potential of tri-lineage differentiation and to investigate the potential of tenogenic differentiation of human tonsil derived MSCs (T-MSCs), bone marrow derived MSCs (BM-MSCs), and adipose tissue derived MSCs (AD-MSCs). Methods: Each tissue was obtained from 8 patients. After isolation of MSCs, flow cytometry analysis was used to characterize the phenotypes of the MSCs. Differentiation capacity to adipo-, osteo-, and chondrocytes were induced by culturing each MSCs for 3 weeks in commercially available media. Each MSCs was treated with 5ng/ml and 10ng/ml of TGF-ß3 with vehicle control. Results: Immunophenotypic surface marker analysis of BM-MSC, AD-MSC, and TMSCs revealed that these MSCs expressed a typical MSCs. mRNA expression levels of the markers for tri-lineage differentiation were significantly lower in TMSC than other MSCs. The tenogenic transcription factor, scleraxis, showed a statistically significant increase in all MSCs differentiation groups except for the 7th day TMSC differentiation group (Figure). Gene expression of tenascin-C, an ECM glycoprotein, was specifically expressed in the T-MSC differentiation group at 14 days (Figure). Comparing the ratio of collagen 1 to collagen 3 genes, the BM-MSC showed a decrease in the ratio on days 3 and 7 unlike AD-MSCs and TMSCs. Only TMSC showed a significant increase in the ratio compared with other MSCs on the 14th day. Conclusion: The tonsil-MSC has low fat, bone and cartilage differentiation potential and has excellent tendon-specific differentiation potential, thus being highly useful as a tendon-tailored cell therapy agent.

Su1180 - The Efficacy of Tonsil-Derived Mesenchymal Stem Cells Conditioned Medium in Chronic Colitis Model

Su1180 - The Efficacy of Tonsil-Derived Mesenchymal Stem Cells Conditioned Medium in Chronic Colitis Model

Regenerative potential of tonsil mesenchymal stem cells on surgical cutaneous defect

As tissue engineering and regenerative medicine have evolved recently, stem cell therapy has been investigated in the field of impaired wound healing. Several studies have reported that mesenchymal stem cells derived from various tissues including bone marrow and adipose tissue can exert the regenerative efficacy in the wound healing. Previously, we have demonstrated the isolation and characterization of tonsil-derived mesenchymal stem cells (TMSCs) with excellent proliferative property. In the present study, we aimed to evaluate the regenerative efficacy of TMSCs in the wound healing process. Two distinct cutaneous surgical defects were generated in the dorsum of mice. Each wound was treated with TMSCs or phosphate-buffered saline (PBS), respectively. After sacrifice, the skin and subcutaneous tissues around the surgical defect were harvested and assessed for inflammation, re-epithelialization, dermal regeneration, and granulation tissue formation. The administration of TMSCs into wound beds significantly promoted the repair of surgical defects in mice. Especially, TMSCs efficiently contributed to the attenuation of excessive inflammation in the surgical lesion, as well as the augmentation of epidermal and dermal regeneration. To elucidate the underlying mechanisms, TMSCs were analyzed for their potency in immunomodulatory ability on immune cells, stimulatory effect on the proliferation of keratinocytes, and fibroblasts, as well as the regulation of fibroblast differentiation. TMSCs inhibited the non-specific or T-cell-specific proliferation of peripheral blood mononuclear cells, as well as the M1 polarization of macrophage-like cells. Moreover, TMSCs augmented the proliferation of skin-constituting fibroblasts and keratinocytes while they suppressed the differentiation of fibroblasts into myofibroblasts. Taken together, our findings demonstrate the regenerative potential of TMSCs in wound healing process through the regulation on inflammation, proliferation, and remodeling of various skin cells, implying that TMSCs can be a promising alternative for wound repair.

P033 The efficacy of tonsil-derived mesenchymal stem cells conditioned medium in chronic colitis model

P033 The efficacy of tonsil-derived mesenchymal stem cells conditioned medium in chronic colitis model

Coordinating Thermogel for Stem Cell Spheroids and Their Cyto‐Effectiveness

AbstractA polymer (mP) with thermogelling and metal coordinating properties is prepared by pyridine‐dicarboxylate (PDC) connected poly(ethylene glycol)‐poly(propylene glycol)‐poly(ethylene glycol) triblock copolymers. Tonsil‐derived mesenchymal stem cells (TMSCs) are incorporated in the mP hydrogel by increasing the temperature of the cell‐suspended aqueous mP solution to 37 °C. The TMSCs are randomly embedded in the in situ formed hydrogel at first; however, they aggregated to form live cell spheroids on day 7. In contrast, the spheroid formation is blocked in the Fe3+‐incorporating mP thermogel. Compared with the conventional 2D‐cultured stem cells, the stem cell spheroid in the 3D mP culture system exhibits significantly enhanced stemness biomarkers, angiogenic biomarkers, and anti‐inflammatory biomarkers in the growth medium. In addition, the stem cell spheroid exhibits significantly greater biomarker expression for osteogenic, chondrogenic, and adipogenic differentiations than the stem cells cultured in the 2D system in each induction medium. This study suggests that a simple injection of stem cells suspended in the current aqueous mP solution can lead to the spontaneous formation of stem cell spheroids with excellent multipotency and retention in the in situ formed thermogel, and thus opens a direct injectable method for the application of the stem cells at a target site.

Layered Double Hydroxide and Polypeptide Thermogel Nanocomposite System for Chondrogenic Differentiation of Stem Cells.

Stem cell therapy for damaged cartilage suffers from low rates of retention, survival, and differentiation into chondrocytes at the target site. To solve these problems, here we propose a two-dimensional/three-dimensional (2D/3D) nanocomposite system. As a new two-dimensional (2D) material, hexagonal layered double hydroxides (LDHs) with a uniform lateral length of 2-3 μm were prepared by a hydrothermal process. Then, tonsil-derived mesenchymal stem cells (TMSCs), arginylglycylaspartic acid-coated LDHs, and kartogenin (KGN) were incorporated into the gel through the thermal-energy-driven gelation of the system. The cells exhibited a tendency to aggregate in the nanocomposite system. In particular, chondrogenic biomarkers of type II collagen and transcription factor SOX 9 significantly increased at both the mRNA and protein levels in the nanocomposite system, compared to the pure thermogel systems. The inorganic 2D materials increased the rigidity of the matrix, slowed down the release of a soluble factor (KGN), and improved cell-material interactions in the gel. The current 2D/3D nanocomposite system of bioactive LDH/thermogel can be a new platform material overcoming drawbacks of hydrogel-based 3D cell culture systems and is eventually expected to be applied as an injectable stem cell therapy.

RNA sequencing reveals a transcriptomic portrait of human mesenchymal stem cells from bone marrow, adipose tissue, and palatine tonsils

Human mesenchymal stem cells (MSCs) are adult multipotent cells that have plasticity and inhabit the stroma of diverse tissues. The potential utility of MSCs has been heavily investigated in the fields of regenerative medicine and cell therapy. However, MSCs represent diverse populations that may depend on the tissue of origin. Thus, the ability to identify specific MSC populations has remained difficult. Using RNA sequencing, we analyzed the whole transcriptomes of bone marrow-derived MSCs (BMs), adipose tissue-derived MSCs (AMs), and tonsil-derived MSCs (TMs). We categorized highly regulated genes from these MSC groups according to functional gene ontology (GO) classification. AMs and TMs showed higher expression of genes encoding proteins that function in protein binding, growth factor, or cytokine activity in extracellular compartments than BMs. Interestingly, TM were highly enriched for genes coding extracellular, protein-binding proteins compared with AMs. Functional Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis also showed differentially enriched signaling pathways between the three MSC groups. Further, we confirmed surface antigens expressed in common and in a tissue-specific manner on BMs, AMs, and TMs by flow cytometry analysis. This study provides comprehensive characteristics of MSCs derived from different tissues to better understand their cellular and molecular biology.

Preventive effects of tonsil-derived mesenchymal stem cells on osteoradionecrosis in a rat model.

The purpose of this study was to investigate the effects of tonsil-derived mesenchymal stem cells (MSCs) on osteoradionecrosis (ORN). We generated a mandibular ORN rat model using a combination of 20-Gy single-dose irradiation and tooth extraction. Study groups were negative control (tooth extraction only), ORN group (irradiation, tooth extraction), Matrigel-1 group (Matrigel; BD Biosciences, San Jose, CA; irradiation, Matrigel application immediately after tooth extraction), tonsil-derived MSC-1 group (irradiation, tonsil-derived MSC application immediately after tooth extraction), Matrigel-4 group (irradiation, Matrigel application 4 weeks after tooth extraction), and tonsil-derived MSC-4 group (irradiation, tonsil-derived MSC application 4 weeks after tooth extraction). Bone mineral density was significantly lower in the ORN group than in the negative control group. The tonsil-derived MSC-1 group showed significantly higher bone mineral density than did the ORN and tonsil-derived MSC-4 groups. A single 20-Gy dose of irradiation combined with tooth extraction successfully generated ORN in the rat model. The tonsil-derived MSCs can be effective for bone regeneration in ORN, particularly when applied immediately after dentoalveolar trauma or surgery.

Hepatogenic Supported Differentiation of Mesenchymal Stem Cells in a Lactobionic Acid-Conjugated Thermogel.

To investigate the effect of receptor substrate of target cells on stem cell differentiation, lactobionic acid-conjugated poly[(propylene glycol)-b-(ethylene glycol)-b-(propylene glycol)]-poly(l-alanine) (LB-PLX-PA) was synthesized, and then thermogelling systems consisting of LB-PLX-PA and PLX-PA in a ratio of 0/100 (LB-0), 5/95 (LB-5), and 20/80 (LB-20) were constructed as an injectable three-dimensional scaffold toward hepatogenic differentiation of tonsil-derived mesenchymal stem cells (TMSCs). Modulus of LB-0, LB-5, and LB-20 increased to 500-800 Pa at 37 °C (gel) due to the heat induced sol-to-gel transition of the systems during which TMSCs were incorporated into the gel. Based on biomarker expressions and hepatic biofunctions of the differentiated cells, the receptor substrate (LB)-conjugated bioactive thermogel provides compatible microenvironments for the differentiated cells, and thus gives pronounced positive results on the differentiation of the stem cells into target cells during three-dimensional culture, compared with a passive thermogel.

Mesenchymal stem cells inhibit RANK-RANKL interactions between osteoclasts and Th17 cells via osteoprotegerin activity.

Th17 cells play a critical role in several autoimmune diseases, including psoriasis and psoriatic arthritis (PsA). Psoriasis is a chronic inflammatory skin disease associated with systemic inflammation and comorbidities, such as PsA. PsA develops in nearly 70% of patients with psoriasis, and osteoclasts associated bone erosion is a hallmark of the disease. Thus far, the effect of Th17 cells on osteoclastogenesis via direct cell-to-cell interactions is less understood. In this study, we observed that Th17 cells directly promote osteoclast differentiation and maturation via expression of receptor activator of nuclear factor-κ β ligand (RANKL) in vitro. We investigated the impact of conditioned medium obtained from human palatine tonsil-derived mesenchymal stem cells (T-CM) on the interactions between osteoclasts and Th17 cells. T-CM effectively blunted the RANK-RANKL interaction between the osteoclast precursor cell line RAW 264.7 and Th17 cells via osteoprotegerin (OPG) activity. The frequency of tartrate-resistant acid phosphatase (TRAP)-positive cells in the bone marrow of an imiquimod (IMQ)-induced psoriasis mouse model was decreased following T-CM injection. Therefore, our data provide novel insight into the therapeutic potential of tonsil-derived mesenchymal stem cell-mediated therapy (via OPG production) for the treatment of pathophysiologic processes induced by osteoclasts under chronic inflammatory conditions such as psoriasis.

Enhanced Osteogenic Commitment of Human Mesenchymal Stem Cells on Polyethylene Glycol-Based Cryogel with Graphene Oxide Substrate.

Graphene oxide (GO) is considered a comparatively recent biomaterial with enormous potential because of its nontoxicity, high dispersity, and enhanced interaction with biomolecules. These characteristics of GO can promote the interactions between the substrates and cell surfaces. In this study, we incorporated GO in a cryogel-based scaffold system to observe their influence on the osteogenic responses of human tonsil-derived mesenchymal stem cells (hTMSCs). Compared to polyethylene glycol (PEG)-based cryogel scaffold, GO-embedded PEG-based (PEGDA-GO) cryogels not only showed improved cell attachment and focal adhesion kinase (FAK) signaling activation but also enhanced cell viability. Taken together, we demonstrated that PEGDA-GO cryogels can stimulate osteogenic differentiation under an osteoinductive condition and enhance osteogenic phenotypes compared to the control group. In summary, we demonstrate that GO embedded in cryogels system is an effective biofunctionalizing scaffold to control osteogenic commitment of stem cells.

Tenogenic differentiation of tonsil-derived mesenchymal stem cells

Category: Basic Sciences/Biologics Introduction/Purpose: Human palatine tonsil-derived mesenchymal stem cells (T-MSCs) are known to be a new source of progenitor cells. Using waste tissue after tonsillectomy as a cell provider can be the biggest benefit of T-MSCs, compared with other stem cells. The purpose of this study was to investigate tenogenic differentiation of T-MSCs and to access the differential effects of TGF-ß3 on the tenogenesis of T-MSCs. Methods: Human tonsil was obtained after tonsillectomy.Using a cytometric analysis, we were able to find that the T-MSCs had typical mesenchymal stem cell markers: positive for CD73, CD90 and CD105, and negative for CD14, CD34 and CD45. Using a Transforming growth factor beta 3 (TGF-ß3), the expressions of tenocyte-specific genes and proteins, such as Collagen type I (COL1), Tenomodulin (TNMD), and Scleraxis (SCX), were measured by a quantitative polymerase chain reaction (PCR) assay, immunofluorescence staining, immunohistochemistry and Western blot analysis. Results: Quantitative PCR assay showed that TGF-ß3 significantly increased the expressions of tenocyte lineage marker genes, including Collagen type I (COL1), Tenomodulin (TNMD), and Scleraxis (SCX), at a 3-day treatment, compared with control. However, these increases were not found at long-term exposures (7 or 10 days), except that TNMD expression was maintained at 50 ng/ml at 7-day exposure to TGF-ß3 (Fig). Like genes, the protein expression levels of COL1, TNMD, and SCX were also induced in TGF-ß3-treated T-MSCsin 3-day treatments. Moreover, the protein levels were maintained for 10 days, as evidenced by immunofluorescence staining, immunohistochemistry and Western blot analyses. Conclusion: This study demonstrated that T-MSCs in tenogenic stimulation with TGF-ß3 have high tenogenic differentiation potential.

Optimization of tenocyte lineage-related factors from tonsil-derived mesenchymal stem cells using design of experiments

Optimization of tenocyte lineage-related factors from tonsil-derived mesenchymal stem cells using design of experiments

Therapeutic potential of tonsil-derived mesenchymal stem cells in dextran sulfate sodium-induced experimental murine colitis.

The therapeutic potential of tonsil-derived mesenchymal stem cells (TMSC) prepared from human tonsillar tissue has been studied in animal models for several diseases such as hepatic injury, hypoparathyroidism, diabetes and muscle dystrophy. In this study, we examined the therapeutic effects of TMSC in a dextran sulfate sodium (DSS)-induced colitis model. TMSC were injected in DSS-induced colitis mice via intraperitoneal injection twice (TMSC[x2]) or four times (TMSC[x4]). Control mice were injected with either phosphate-buffered saline or human embryonic kidney 293 cells. Body weight, stool condition and disease activity index (DAI) were examined daily. Colon length, histologic grading, and mRNA expression of pro-inflammatory cytokines, interleukin 1β (IL-1β), IL-6, IL-17 and tumor necrosis factor α, and anti-inflammatory cytokines, IL-10, IL-11 and IL-13, were also measured. Our results showed a significant improvement in survival rates and body weight gain in colitis mice injected with TMSC[x2] or TMSC[x4]. Injection with TMSC also significantly decreased DAI scores throughout the experimental period; at the end of experiment, almost complete reversal of DAI scores to normal was found in colitis mice treated with TMSC[x4]. Colon length was also significantly recovered in colitis mice treated with TMSC[x4]. However, histopathological alterations induced by DSS treatment were not apparently improved by injection with TMSC. Finally, treatment with TMSC[x4] significantly reversed the mRNA levels of IL-1β and IL-6, although expression of all pro-inflammatory cytokines tested was induced in colitis mice. Under our experimental conditions, however, no apparent alterations in the mRNA levels of all the anti-inflammatory cytokines tested were found. In conclusion, our findings demonstrate that multiple injections with TMSC produced a therapeutic effect in a mouse model of DSS-induced colitis.

Application of differentiated human tonsil-derived stem cells to trembler-J mice.

Mesenchymal stem cells (MSCs) can differentiate into various cell types. In this study we investigated the potential of human tonsil-derived MSCs (T-MSCs) for neuromuscular regeneration in trembler-J (Tr-J) mice, a model for Charcot-Marie-Tooth disease type 1A (CMT1A). T-MSCs differentiated toward skeletal myocytes with increased expression of skeletal muscle-related markers (including troponin I type 1, and myogenin), and the formation of myotubes in vitro. In-situ transplantation of T-MSC-derived myocytes (T-MSC myocytes) into the gastrocnemius muscle in Tr-J mice enhanced motor function, with recovery of compound muscle action potential amplitudes. Morphology of the sciatic nerve and skeletal muscle recovered without the formation of teratomas, and the expression levels of nerve growth factor and glial-cell-line-derived neurotrophic factor were increased significantly in T-MSC myocytes compared with T-MSCs in vitro. Transplantation of T-MSC myocytes could enable neuromuscular regeneration in patients with CMT1A. Muscle Nerve 57: 478-486, 2018.

Intracellular Remodeling and Accumulation of Aberrant Lysosomes in Differentiation of Tonsil-Derived Mesenchymal Stem Cells into Parathyroid-Like Cells.

Differentiation of mesenchymal stem cells (MSC) into a variety of cell lineages such as adipocytes, osteocytes, and chondrocytes is often accompanied up-regulation of autophagy. In our study, we demonstrated that the expression of autophagy-associated proteins (p-Beclin 1, LC3A, LC3B, p-AMPK, p-mTOR and ATG3, ATG7, and ATG12-5) over a period of time was hardly distinguishable from control tonsil-derived MSC (TMSC). Despite the unnoticeable difference in autophagy activation between differentiated TMSC (dTMSC) and the control (cTMSC), we reported significant changes in intracellular compositions in differentiated TMSC into functional parathyroid-like cells secreting parathyroid hormone (PTH). By using transmission electron microscopy (TEM), we observed accumulation of multivesicular bodies (MVB) comprising small, degraded compartments densely accumulated as dark granular or amorphous clumps, multilamellar bodies and lipid droplets in dTMSC. However, no such structures were found in cTMSC. These results suggest that differentiation of TMSC into parathyroid-like cells producing PTH hormone is hardly dependent on autophagy activation in the beginning of our conditions. Furthermore, our results of intracellular remodeling and accumulated endo-lysosomal storage bodies in the later stages of TMSC differentiation present a possible role of the structures in PTH secretion.