Oat (Avena sativa L.) Sprouts Restore Skin Barrier Function by Modulating the Expression of the Epidermal Differentiation Complex in Models of Skin Irritation.

Oats (Avena sativa L.) are used as therapeutic plants, particularly in dermatology. Despite numerous studies on their skin moisturization, anti-inflammation, and antioxidation effects, the precise molecular mechanisms of these effects are only partially understood. In this study, the efficacy of oat sprouts in the treatment of allergic contact dermatitis (ACD) was investigated, and their specific phytoconstituents and exact mechanisms of action were identified. In the in vivo ACD model, by stimulating the mitogen-activated protein kinase signaling pathway, oat sprouts increased the expression levels of proteins associated with skin barrier formation, which are produced during the differentiation of keratinocytes. In addition, in a lipopolysaccharide-induced skin irritation model using HaCaT, steroidal saponins (avenacoside B and 26-deglucoavenacoside B) and a flavonoid (isovitexin-2-o-arabinoside) of oat sprouts regulated the genetic expression of the same proteins located on the adjacent locus of human chromosomes known as the epidermal differentiation complex (EDC). Furthermore, oat sprouts showed immunomodulatory functions. These findings suggest the potential for expanding the use of oat sprouts as a treatment option for various diseases characterized by skin barrier disruption.

Adjuvant therapy with imatinib for an incompletely resected multilobular tumour of bone in a dog.

A 5-year-old neutered male Shiba Inu dog presented with a history of oral bleeding, dysphagia, and depression for 3 weeks. The physical examination revealed a firm mass in the right caudal palatal region along the level of PM4-M2. On the computed tomography, the mass was round-to-oval in shape and 22 mm × 30 mm × 15 mm in size. The mass contained multiple bone attenuated materials with a palatal bone lysis of 4 mm × 6 mm. A complete resection of the mass was proposed; however, the owner declined due to the risk of complications associated with the radical surgery. Therefore, a palliative resection and biopsy of the mass were performed. On the histological examination, the mass was diagnosed as grade 2 multilobular tumour of bone (MTB). Since the mass was incompletely resected, adjuvant therapy was pursued along with targeted therapy using a tyrosine kinase inhibitor. The tumour cells showed overexpression of the receptor of tyrosine kinase for c-KIT, PDGFR-α, PDGFR-ß, and FGFR1 compared to normal tissue cells. Additionally, the cytotoxic effect of imatinib on the MTB cells was confirmed in vitro. Four weeks postoperatively, the administration of imatinib and carprofen was initiated and continued for 259 days. The patient maintained a good functional outcome for 306 days after the initial presentation.

Regulation of Adipogenesis Through Differential Modulation of ROS and Kinase Signaling Pathways by 3,4'-Dihydroxyflavone Treatment.

Studies on adipogenesis may be important for regulating human and/or animal obesity, which causes several complications such as, type II diabetes, hypertension, and cardiovascular disease, thus giving rise to increased economic burden in many countries. Previous reports revealed that various flavonoids have anti-apoptotic, antioxidant, and cell differentiation-regulating activities with a number of physiological benefits, including protection from cardiovascular disease, cancers, and oxidative stress. As we found that the hydroxylation patterns of the flavonoid B ring are known to play a critical role in their function, we screened several flavonoids containing different numbers and positions of OH substitutions in B ring for their modulatory property on adipogenesis. In this study, we revealed the anti-adipogenic activity of the naturally derived flavonoid, 3,4'-dihydroxyflavone (3,4'-DHF) in murine 3T3-L1 pre-adipocytes and equine adipose-derived stromal cells (eADSCs). We found that treatment with 3,4'-dihydroxyflavone (3,4'-DHF) led to decreased expression of adipogenic markers and lipid deposition with differential modulation of ROS and kinase signaling pathways. Regulation of ROS generation through the differential modulation of ROS-regulating gene expression was revealed to have an important role in the suppression of adipogenesis and increase of osteogenesis in eADSCs following 3,4'-DHF treatment. These results suggest that the flavonoid 3,4'-DHF can be used to regulate adipogenesis in ADSCs, which has potential therapeutic application in regenerative medicine or health care for humans and many sport or companion animals. J. Cell. Biochem. 118: 1065-1077, 2017. © 2016 Wiley Periodicals, Inc.

Platelet-Rich Plasma Increases the Levels of Catabolic Molecules and Cellular Dedifferentiation in the Meniscus of a Rabbit Model.

Despite the susceptibility to frequent intrinsic and extrinsic injuries, especially in the inner zone, the meniscus does not heal spontaneously owing to its poor vascularity. In this study, the effect of platelet-rich plasma (PRP), containing various growth factors, on meniscal mechanisms was examined under normal and post-traumatic inflammatory conditions. Isolated primary meniscal cells of New Zealand white (NZW) rabbits were incubated for 3, 10, 14 and 21 days with PRP(-), 10% PRP (PRP(+)), IL(+) or IL(+)PRP(+). The meniscal cells were collected and examined using reverse-transcription polymerase chain reaction (RT-PCR). Culture media were examined by immunoblot analyses for matrix metalloproteinases (MMP) catabolic molecules. PRP containing growth factors improved the cellular viability of meniscal cells in a concentration-dependent manner at Days 1, 4 and 7. However, based on RT-PCR, meniscal cells demonstrated dedifferentiation, along with an increase in type I collagen in the PRP(+) and in IL(+)PRP(+). In PRP(+), the aggrecan expression levels were lower than in the PRP(-) until Day 21. The protein levels of MMP-1 and MMP-3 were higher in each PRP group, i.e., PRP(+) and IL(+)PRP(+), at each culture time. A reproducible 2-mm circular defect on the meniscus of NZW rabbit was used to implant fibrin glue (control) or PRP in vivo. After eight weeks, the lesions in the control and PRP groups were occupied with fibrous tissue, but not with meniscal cells. This study shows that PRP treatment of the meniscus results in an increase of catabolic molecules, especially those related to IL-1α-induced inflammation, and that PRP treatment for an in vivo meniscus injury accelerates fibrosis, instead of meniscal cartilage.

Stimulation of cannabinoid receptors by using Rubus coreanus extracts to control osteoporosis in aged male rats.

A substantial proportion of men with prostatic disease have an increased risk of bone loss. In the present study, we investigated the effects of Rubus coreanus Miquel (RCM) extracts on osteoporosis that occurs with N-methyl-N-nitrosourea (MNU)-induced prostatic hyperplasia. The rats used in this study were categorized into groups of healthy controls, rats treated with MNU, and rats treated with MNU and RCM. The rats were sacrificed after 10 weeks of RCM treatment, after which ultrasonography, serum biochemical tests, histopathological examinations, immunohistochemical analysis, and semi-quantitative reverse-transcription polymerase chain reaction analysis were performed. There were no marked differences in body weight gain and the size and weight of the prostate gland between the MNU group and the MNU and RCM group. However, treatment with RCM inhibited osteoclastic osteolysis and reduced dysplastic progress in the prostate gland, as observed by histopathological evaluation and by analyzing changes in the levels of bone regulatory factors. In addition, the group treated with MNU and RCM had higher expression levels of cannabinoid receptors-1, -2, and osteoprotegerin. These results indicate that the anti-osteoporotic effect of RCM in prostatic hyperplasia is attributable to the cannabinoid receptor-related upregulation of osteoblastogenesis and inhibition of prostatic hyperplasia. The results of the present study suggest that treatment with RCM may benefit osteoporotic patients with prostatic disease by simultaneously altering the activation of osteoblasts and osteoclasts.

Changes of the Cytoplasmic Proteome in Response to Alcoholic Hepatotoxicity in Rats.

Proteomic analyses have already been used in a number of hepatological studies and provide important information. However, few reports have focused on changes in the cytoplasmic proteome. The present study therefore aimed to evaluate changes in cytoplasmic proteome of rats in response to alcoholic hepatotoxicity. Rats were fed a Liber-DeCarli liquid diet containing ethanol for four weeks. Cytoplasmic proteins except mitochondrial proteins from the livers of these animals were investigated using two-dimensional gel electrophoresis and mass spectrometry. Alcohol induced a decrease in body weight gain and an increase in alanine transaminase (ALT), cholesterol, and phospholipid levels. Histopathological observations revealed hepatic damage characterized by necrosis and fatty change in alcohol-treated group at week 2, which continues until week 4. Our proteomic analysis revealed that 25 proteins were differentially expressed in the ethanol-fed group. Of these, 12 cytoplasmic proteins are being reported for the first time. Taken together, our results provide further insights into the disease mechanism and therapeutic information of alcoholic liver disease.

Abnormal changes in both mandibular salivary glands in a dog: Non-mineral radiopaque sialoliths.

A 10-year-old Maltese dog was presented with a firm mass on the left side of his neck. Physical examination confirmed a firm mass in the left and a submandibular swelling in the right cervical region. Sialolithiasis and associated sialocele in both mandibular salivary glands were suspected and bilateral sialoadenectomy was performed. The stones were identified as non-mineral sialoliths.

Changements anormaux dans les deux glandes salivaires mandibulaires chez un chien : sialolithes radiopaques non minéraux. Un chien Maltais âgé de 10 ans a été présenté avec une masse ferme du côté droit du cou. L'examen physique a confirmé une masse ferme dans l'enflure gauche et sous-mandibulaire dans la région cervicale droite. La sialolithiase et une sialocèle connexe dans les deux glandes salivaires mandibulaires ont été suspectées et une sialoadénectomie bilatérale a été réalisée. Les pierres ont été identifiées comme des sialolithes non minéraux.(Traduit par Isabelle Vallières).

Primary lymphoma of the uterine horn in a Lhasa Apso dog.

Primary lymphomas of the canine female genital tract are uncommon tumours. A 9-year-old intact female Lhasa Apso dog presenting with a closed pyometra underwent an ovariohysterectomy (OHE), and the hyperplastic uterine horn along with multiple follicular cysts on the right ovary was examined by histological analysis. Severe infiltration of medium-sized lymphocytes with strong positive immunoreactivity for CD79a and numerous anaplastic features was detected in the unilateral uterine horn, and the dog was diagnosed as having extranodal marginal zone B-cell lymphoma (MZBCL). The present case reports an extremely rare occurrence of primary lymphoma involving the uterine horn in a dog and describes histological characteristics of the tumour for definite diagnosis.

Time-sequential modulation in expression of growth factors from platelet-rich plasma (PRP) on the chondrocyte cultures.

Platelets are involved in hemostasis, wound healing, and tumor growth. Autologous blood products are commonly used to facilitate healing in a variety of clinical surgery applications. Recently, it was shown that platelet-rich plasma (PRP) has more specific growth factors that participate in the healing process. This study investigated the expression of PRP growth factors and evaluated their potential role in the cartilage regeneration using primary isolated chondrocytes. PRP obtained from New Zealand White rabbit by low speed centrifugation. Extracted PRPs contained 6-10 × 10(6) platelet/µl and concentration of platelets was slightly variable. Primary isolated chondrocytes from the same rabbits were cultured and treated with 0.1-20% PRP. The cells were collected and examined by reverse transcription-polymerase chain reaction and cytochemical staining. The expression of sex determining region Y-box 9, transforming growth factor-beta, vascular endothelial growth factor, and chondromdulin-I was increased in chondrocyte cultures with 10% PRP by time-dependent manner. To maintain the integrity of the cartilage, the proteoglycan contents were also up-regulated from the mRNA of aggrecan and positive Safranin-O staining in PRP concentration- and time-dependent manner. PRP provides crucial growth factors related to chondrocyte proliferation and differentiation through time-sequential modulation. Controlled in vivo trials for cartilage regeneration are needed.

Anterior cruciate ligament reconstruction in a rabbit model using canine small intestinal submucosa and autologous platelet-rich plasma.

BACKGROUND: The bone-ligament interface is the main point of failure after anterior cruciate ligament reconstruction. Synthetic ligament materials have problems such as a greater failure rate of the bone-ligament insertion than autografts. Small intestinal submucosa (SIS) is a biologic scaffold that has been used to repair musculoskeletal tissue and has been shown to promote cell migration and enhance collagen fiber regeneration. Autologous platelet-rich plasma (PRP) has also been investigated as a potential promoter of tendon healing. We investigated SIS and PRP as biomaterials that might strengthen the bone-tunnel interface and improve tendon structure formation. METHODS: Anterior cruciate ligament grafts were formed of braid-twist canine SIS. These canine SIS ligament grafts were used for anterior cruciate ligament reconstruction in 20 New Zealand white rabbits. The rabbits were divided into 2 treatment groups. In 1 group (SIS group; n = 10), we only implanted the canine SIS grafts. In the second group (PRP group; n = 10), we applied autologous PRP to the surgical area after implantation of canine SIS grafts. We determined the cytokine level of the autologous PRP using a transforming growth factor-ß1 enzyme-linked immunosorbent assay kit. At 1 and 4 wk after surgery, magnetic resonance imaging was performed to evaluate the grafts. The femur-graft-tibia complex was assessed histologically and biomechanically at 8 wk after surgery. RESULTS: At 1 wk after surgery, the magnetic resonance imaging scans of the PRP group showed high signal-intensity lesions. In biomechanical tests, the SIS group had a significantly greater maximum load, maximum stress, and ultimate load and strain than the PRP group. The histologic findings of the PRP group revealed a greater cellular response, fibrotic tissue regeneration around the graft, broad chondrocyte cell infiltration, and collagen fibers that were loosely attached to the bone. CONCLUSIONS: The PRP group had significantly lower tension load values than the SIS group, and there was greater cellular response in a broad area around the grafts of the rabbits in the PRP group compared with those in the SIS group. The early inflammatory responses around the canine SIS grafts in the PRP group and the altered cytokine or growth factor concentration in the intra-articular capsule of the rabbits in PRP group might explain their relatively low tensile strength results.

Tracheal replacement with fresh and cryopreserved aortic allograft in adult dog.

BACKGROUND: Many reports have described tracheal replacement using aortic allografts, with varying results and minimal understanding of the mechanism of tracheal regeneration. The present study attempted tracheal regeneration in adult dogs using fresh aortic allografts (FAA) and cryopreserved aortic allografts (CAA). MATERIALS AND METHODS: Twelve adult beagles underwent tracheal resection and were transplanted with FAA (n = 5) or CAA (n = 7). Animals were followed-up with serial radiography and magnetic resonance imaging, and were euthanized at predetermined times up to 16 mo post-surgery. RESULTS: There were no procedural deaths, but two animals died due to stent migration. Stent migration occurred in seven of the 12 animals. Evidence of regeneration of tracheal epithelium was observed in the surviving animals, with the transformation of squamous metaplasia to mucociliary epithelium being time-dependent. Islet of cartilage were observed in animals after 6 mo, but ring-like cartilage structures were absent, even after 16 mo. During autopsy, axial graft contractions up to 68% were observed. Serial radiographs show that most of the contraction occurred within 1 mo. The results of the MRI showed that the graft area was strongly enhanced for up to 2 mo, but was clearly reduced after 3 mo. CONCLUSIONS: Tracheal replacement in adult dogs using FAA or CAA is feasible. However, immaturity of the neotracheal cartilage did not allow the tissue to function as native tracheal tissue. Prolonged stenting should be considered in adult if the procedure is to be clinically contemplated.

Case Report: Articular Gout in Four Dogs and One Cat.

Background: There is widespread prejudice in veterinary medicine that gout does not occur in non-human mammalians. However, we recently discovered monosodium urate crystals in the synovial fluid obtained from a few dogs and a cat. Since it is the definitive and gold standard to diagnose gout, we report these cases as newly emerging diseases in companion animals. Case Presentation: Four dogs and one cat were presented at our hospital because of lameness due to an unknown cause. Even after the routine examinations, including radiographic imaging, laboratory examination, and arthrocentesis, we were unable to find a clear cause of polyarthritis. However, we later discovered monosodium urate crystals in the synovial fluid of the animals, confirmed by polarized microscopy. In one of the two dogs treated with immunosuppressants, the disease relapsed, and the other did not show any symptoms for 3 months. The other two dogs were treated with xanthine oxidase inhibitor, where one died, and the other did not show any symptoms for 3 years. The cat was treated with drainage and intra-articular dexamethasone injection, but the disease recurred after 6 months. Conclusion: This is the first report to confirm that articular gout can occur in dogs and cats. Care must be taken not to neglect needle-shaped materials in the synovial fluid. Gout should also be included in the differential diagnosis of arthritis and further research is needed in these animals.

Changes in metabolites with harvest times of seedlings of various Korean oat (Avena sativa L.) cultivars and their neuraminidase inhibitory effects.

Oats and their seeds, stems, and leaves are approved for use as safe food ingredients. Oat seedlings are environmentally friendly and are becoming increasingly popular as they provide several health benefits. We used the UPLC-CAD to quantitatively analyze isolated compounds (1-11) between 15 cultivars of oat seedlings and their harvest time. Maximum average amount of total contents of isolated compounds was observed after the harvest time of 5 days (4711.3 mg/100 g), while the minimum was observed after the harvest time of 7 days (4184.8 mg/100 g). We demonstrated that all isolated compounds (1-11) showed neuraminidase inhibitory effects, with 6 and 7 being the most active with IC50 values of 3.7 and 20.5 µM, respectively. High content of compounds 6 and 7 was observed (2306.6 mg/100 g) in the Dahan cultivar at 9 days, indicating potential good cultivars with a high content of active compounds and neuraminidase inhibition activity.

Effect of Decellularized Extracellular Matrix Bioscaffolds Derived from Fibroblasts on Skin Wound Healing and Remodeling.

The mammalian tissue extracellular matrix (ECM) has been used as a scaffold to facilitate the repair and reconstruction of numerous tissues. However, the material properties of decellularized ECM (dECM) from in vitro cell cultures and the effect of these properties on wound remodeling remain unclear. To elucidate its biological activity, we extracted dECM from human lung fibroblasts, fabricated it into a patch, and applied it to a full-thickness skin wound. The fibroblast-derived dECM (fdECM) maintained the content of collagen Ⅰ, collagen Ⅳ, and elastin, and the extraction process did not damage its critical growth factors. The fdECM-conjugated collagen patch (COL-fdECM) facilitated wound contraction and angiogenesis in the proliferative phase when applied to the in vivo full-thickness skin wound model. Moreover, the COL-fdECM treated wound showed increased regeneration of the epidermal barrier function, mature collagen, hair follicle, and subepidermal nerve plexus, suggesting qualitative skin remodeling. This therapeutic efficacy was similarly observed when applied to the diabetic ulcer model. fdECM was shown to help remodel the tissue by regulating fibroblast growth factors, matrix metalloproteinases, and tissue inhibitors of metalloproteinases via the p38 and ERK signaling pathways in an in vitro experiment for understanding the underlying mechanism. These results provide a biological basis for cell-derived ECM as a multi-functional biomaterial applicable to various diseases.

Keratin-mediated hair growth and its underlying biological mechanism.

Here we show that intradermal injection of keratin promotes hair growth in mice, which results from extracellular interaction of keratin with hair forming cells. Extracellular application of keratin induces condensation of dermal papilla cells and the generation of a P-cadherin-expressing cell population (hair germ) from outer root sheath cells via keratin-mediated microenvironmental changes. Exogenous keratin-mediated hair growth is reflected by the finding that keratin exposure from transforming growth factor beta 2 (TGFß2)-induced apoptotic outer root sheath cells appears to be critical for dermal papilla cell condensation and P-cadherin-expressing hair germ formation. Immunodepletion or downregulation of keratin released from or expressed in TGFß2-induced apoptotic outer root sheath cells negatively influences dermal papilla cell condensation and hair germ formation. Our pilot study provides an evidence on initiating hair regeneration and insight into the biological function of keratin exposed from apoptotic epithelial cells in tissue regeneration and development.

Immunophenotyping of an Unusual Mixed-Type Extraskeletal Osteosarcoma in a Dog.

A 6-year-old female Maltese dog presented with a cervical mass without pain. The tumor was surrounded by a thick fibrous tissue and consisted of an osteoid matrix with osteoblasts and two distinct areas: a mesenchymal cell-rich lesion with numerous multinucleated giant cells and a chondroid matrix-rich lesion. The tumor cells exhibited heterogeneous protein expression, including a positive expression of vimentin, cytokeratin, RANKL, CRLR, SOX9, and collagen 2, and was diagnosed as extraskeletal osteosarcoma. Despite its malignancy, the dog showed no sign of recurrence or metastasis three months after the resection. Further analysis of the tumor cells revealed a high expression of proliferation- and metastasis-related biomarkers in the absence of angiogenesis-related biomarkers, suggesting that the lack of angiogenesis and the elevated tumor-associated fibrosis resulted in a hypoxic tumor microenvironment and prevented metastasis.

Non-thermal plasma promotes hair growth by improving the inter-follicular macroenvironment.

Non-thermal plasma (NTP) is widely used in the disinfection and surface modification of biomaterials. NTP treatment can regenerate and improve skin function; however, its effectiveness on hair follicle (HF) growth and its underlying mechanisms need to be elucidated. Herein, we propose an air-based NTP treatment, which generates exogenous nitric oxide (eNO), as a therapeutic strategy for hair growth. The topical application of air-based NTP generates large amounts of eNO, which can be directly detected using a microelectrode NO sensor, in the dermis of mouse dorsal skin. Additionally, NTP-induced eNO has no cytotoxicity in normal human skin cells and promotes hair growth by increasing capillary tube formation, cellular proliferation, and hair/angiogenesis-related protein expression. Furthermore, NTP treatment promotes hair growth with adipogenesis and activation of CD34+CD44+ stem cells and improves the inter-follicular macroenvironment via increased perifollicular vascularity in the mouse hair regrowth model. Given the importance of the hair follicle (HF) cycle ratio (growth vs. regression vs. resting) in diagnosing alopecia, NTP treatment upregulates the stem cell activity of the HF to promote the anagen : catagen : telogen ratio, leading to improved hair growth. We confirmed the upregulation of increasing Wnt/ß-catenin signaling and activation of perifollicular adipose tissue and angiogenesis in HF regeneration. In conclusion, these results show that the eNO from NTP enhances the cellular activities of human skin cells and endothelial cells in vitro and stem cells in vivo, thereby increasing angiogenesis, adipogenesis, and hair growth in the skin dermis. Furthermore, the results of this study suggest that NTP treatment may be a highly efficient alternative in regenerative medicine for achieving enhanced hair growth.

The Effectiveness of Compartmentalized Bone Graft Sponges Made Using Complementary Bone Graft Materials and Succinylated Chitosan Hydrogels.

Bone defects can occur from many causes, including disease or trauma. Bone graft materials (BGMs) have been used to fill damaged areas for the reconstruction of diseased bone tissues since they are cost effective and readily available. However, BGMs quickly disperse around the tissue area, which ultimately leads to it migrating away from the defect after transplantation. We tested chitosan hydrogels as a useful carrier to hold BGMs in the transplantation area. In this study, we synthesized succinylated chitosan (SCS)-based hydrogels with a high decomposition rate and excellent biocompatibility. We confirmed that BGMs were well distributed inside the SCS hydrogel. The SCS-B hydrogel showed a decrease in mechanical properties, such as compressive strength and Young's modulus, as the succinylation rate increased. SCS-B hydrogels also exhibited a high cell growth rate and bone differentiation rate. Moreover, the in vivo results showed that the SCS hydrogel resorbed into the surrounding tissues while maintaining the BGMs in the transplantation area for up to 6 weeks. These data support the idea that SCS hydrogel can be useful as a bioactive drug carrier for a broad range of biomedical applications.

Multiple perianal infundibular follicular cysts in a dog.

This case report describes a 7-year-old male cocker spaniel dog with multiple perianal infundibular follicular cysts. Clinically the dog had moderate anal sacculitis, peri-anal pruritus causing it to 'scoot' and lick the area. On examination of the perianal area, there were over 100 firm, well circumscribed papules, ranged from 0.2 to 0.5 cm in diameter with a central pore, and were found in the perianal region. Alopecia was present in the perianal region. The skin tissue in the perianal region resected surgically was submitted for histological examination. Microscopically, the tissue revealed multiple dilated cysts filled with keratins and the papules corresponded to infundibular follicular cysts. The affected dog showed moderate anal sacculitis. Anal sacculitis commonly causes repeated scooting or licking the area around the anus. Therefore, the multiple follicular cysts in the present case appear to be primarily a sequela to chronic external trauma to the perianal area, probably in response to anal sacculitis. To the best of the authors' knowledge, the present report is the first documented case of multiple perianal infundibular follicular cysts in a dog.

Hypoxia Helps Maintain Nucleus Pulposus Homeostasis by Balancing Autophagy and Apoptosis.

Intervertebral disc degeneration (IVDD) is a common cause of lower back pain. Programmed cell death (PCD) including apoptosis and autophagy is known to play key mechanistic roles in the development of IVDD. We hypothesized that the nucleus pulposus cells that make up the center of the IVD can be affected by aging and environmental oxygen concentration, thus affecting the development of IVDD. Here, we evaluated the phenotype changes and PCD signaling in nucleus pulposus cells in two different oxygen percentages (5% (hypoxia) and 20% (normoxia)) up to serial passage 20. NP cells were isolated from the lumbar discs of rats, and the chondrogenic, autophagic, and apoptotic gene expressions were analyzed during cell culture up to serial passage 20. Hypoxia significantly increased the number of autophagosomes, as determined by monodansylcadaverine staining and transmission electron microscopy. Furthermore, hypoxia triggered the activation of autophagic flux (beclin-1, LC3-II/LC3-I ratio, and SIRT1) with a concomitant decrease in the expression of apoptotic proteins (Bax and caspase-3). Despite injury and age differences, no significant differences were observed between the ex vivo lumbar disc cultures of groups incubated in the hypoxic chamber. Our study provides a better understanding of autophagy- and apoptosis-related senescence in NP cells. These results also provide insight into the effects of aging on NP cells and their PCD levels during aging.