Association between lymph node ratio and survival outcomes in patients with oral squamous cell carcinoma.

BACKGROUND: Recent reports have shown that the Lymph node ratio (LNR) is useful for predicting the prognosis in some cancers, however there are few reports on the usefulness of LNR in predicting the prognosis of oral squamous cell carcinoma (OSCC). The predictive value of LNR for prognosis of OSCC was investigated. MATERIALS AND METHODS: The study included 152 patients with OSCC and histologically confirmed cervical lymph node metastasis who underwent neck dissection. We analyzed the relationship between LNR and overall survival (OS) and recurrence-free survival (RFS) retrospectively in these cases, with the relationship between prognosis and clinicopathological findings also examined. RESULTS: Using a receiver operating characteristics curve, the LNR cutoff value was set at 0.095, categorizing 64 and 88 cases into high LNR (≥ 0.095) and low LNR (< 0.095) groups, respectively. Regarding OS and RFS, the prognosis was significantly worse in the high LNR group compared with the low LNR group. In multivariate analysis, sex, postoperative nodal stage, and LNR merged as independent prognostic factors. CONCLUSION: This study's findings suggest that LNR may represent a prognostic indicator in OSCC with cervical lymph node metastasis.

Imaging Characteristics of Embedded Tooth-Associated Cemento-Osseous Dysplasia by Retrospective Study.

BACKGROUND: Since there are many differential diagnoses for cemento-osseous dysplasia (COD), it is very difficult for dentists to avoid misdiagnosis. In particular, if COD is related to an embedded tooth, differential diagnosis is difficult. However, there have been no reports on the characteristics of the imaging findings of COD associated with embedded teeth. The aim of the present study was to investigate the occurrence and imaging characteristics of cemento-osseous dysplasia (COD) associated with embedded teeth, in order to appropriately diagnose COD with embedded teeth. METHODS: The radiographs with or without histological findings of 225 patients with COD were retrospectively analyzed. A retrospective search through the picture archiving and communication system (PACS) of the Division of Oral and Maxillofacial Radiology of Kyushu Dental University Hospital was performed to identify patients with COD between 2011 and 2022. RESULTS: Fifteen COD-associated embedded mandibular third molars were identified in 13 patients. All 13 patients were asymptomatic. On imaging, COD associated with embedded mandibular third molars appeared as masses that included calcifications around the apex of the tooth. On panoramic tomography, COD showed inconspicuous internal calcification similar to that of odontogenic cysts or simple bone cysts, especially in patients with COD only around the mandibular third molar region. Those with prominent calcification resembled cemento-ossifying fibroma, calcifying epithelial odontogenic tumor, calcifying odontogenic cyst, adenomatoid odontogenic tumor, and so on, as categories of masses that include calcifications on panoramic tomography and computed tomography. CONCLUSIONS: The current investigation is the first to report and analyze the imaging characteristics of COD associated with embedded teeth. It is important to consider the differences between COD and other cystic lesions on panoramic tomography, and the differences between COD and masses that include calcifications on CT.

Fabrication of a Three-Dimensional Spheroid Culture System for Oral Squamous Cell Carcinomas Using a Microfabricated Device.

Cancer stem cells (CSCs) are considered to be responsible for recurrence, metastasis, and resistance to treatment in many types of cancers; therefore, new treatment strategies targeting CSCs are attracting attention. In this study, we fabricated a polyethylene glycol-tagged microwell device that enabled spheroid formation from human oral squamous carcinoma cells. HSC-3 and Ca9-22 cells cultured in the microwell device aggregated and generated a single spheroid per well within 24-48 h. The circular shape and smooth surface of spheroids were maintained for up to five days, and most cells comprising the spheroids were Calcein AM-positive viable cells. Interestingly, the mRNA expression of CSC markers (Cd44, Oct4, Nanog, and Sox2) were significantly higher in the spheroids than in the monolayer cultures. CSC marker-positive cells were observed throughout the spheroids. Moreover, resistance to cisplatin was enhanced in spheroid-cultured cells compared to that in the monolayer-cultured cells. Furthermore, some CSC marker genes were upregulated in HSC-3 and Ca9-22 cells that were outgrown from spheroids. In xenograft model, the tumor growth in the spheroid implantation group was comparable to that in the monolayer culture group. These results suggest that our spheroid culture system may be a high-throughput tool for producing uniform CSCs in large numbers from oral cancer cells.

Horseshoe osteotomy maintains the nasal cavity and function after superior repositioning.

The aim of this prospective cohort study was to compare changes in nasal cavity and function between Le Fort I with and without horseshoe osteotomy after superior repositioning of the maxilla. The patients were divided into 2 groups, a Le Fort I alone (LF alone) group and a combination Le Fort I and horseshoe osteotomy (HS) group. The nasal cavity volume was measured using 3-dimensional computed tomographic images, and nasal resistance was assessed by anterior active mask rhinomanometry. The HS group consisted of 17 patients, and the LF alone group consisted of 15 patients. The magnitude of change in nasal cavity volume was significantly smaller in the HS group than in the LF alone group (p < 0.001), even though the mean amount of superior maxillary movement was considerably larger in the HS group than in the LF alone group (p < 0.001). Mean nasal resistance was significantly smaller postoperatively than preoperatively in the HS group (p < 0.05). Furthermore, the change in nasal resistance was smaller in the HS group than in the LF alone group (p < 0.001). Within the limitations of this study, it seems that horseshoe osteotomy is useful for maintaining the nasal cavity and function after superior repositioning of the maxilla.

Total Hemoglobin Mass Correlates with Peak Oxygen Consumption in Patients with Chronic Stroke.

BACKGROUND: The peak oxygen consumption (V.O2peak) and blood hemoglobin concentration [Hb] are lower in stroke patients than in age-matched healthy subjects. The ability of skeletal muscles to extract oxygen is diminished after stroke. We hypothesized that the oxygen extraction capacity of skeletal muscles in stroke patients depends on [Hb]. To test the hypothesis, we determined the relationship between V.O2peak and total hemoglobin mass (tHb-mass) in stroke patients. METHODS: The subjects were 19 stroke patients (age: 74 ± 2, mean ± SD, 10 males) and 11 age-matched normal subjects (age 76 ± 3, 6 males). Plasma volume (PV) and V.O2peak were measured on the same day. PV was measured using Evans Blue dye dilution method. Blood volume (BV) was calculated from PV and hematocrit, while tHb-mass was estimated from BV and [Hb]. Each subject underwent cardiopulmonary exercise test on a bicycle ergometer using a V.O2peak respiratory gas analyzer. RESULTS: There were no differences in age, height, and weight between the two groups. V.O2peak was lower in stroke patients than in the control. BV and tHb mass were not significantly different between the two groups, but [Hb] was significantly lower in stroke patients. In stroke patients, V.O2peak correlated significantly with tHb-mass (r = 0.497, p < 0.05), but not with BV. CONCLUSION: Our results suggested that low [Hb] seems to contribute to V.O2peak in stroke patients. The significant correlation between tHb-mass and V.O2peak suggested that treatment to improve [Hb] can potentially improve V.O2peak in stroke patients.

Effects of Combination of Functional Electric Stimulation and Robotic Leg Movement Using Dynamic Tilt Table on Walking Characteristics in Post-Stroke Patients with Spastic Hemiplegia: A Randomized Crossover-Controlled Trial.

BACKGROUND: Spastic hemiplegia causes slow and unstable walking in post-stroke patients. Dynamic tilt table with robotic leg movement (DTTRLM) is safe and effective in improving walking. Functional electric stimulation (FES) improves walking speed in post-stroke patients with spastic hemiplegia. The aim of this study was to determine the effects of combined DTTRLM + FES on walking speed compared with DTTRLM alone. METHODS: Twenty post-stroke patients were randomly assigned to receive either a single session of stepping + FES treatment or a single session of stepping alone treatment. After a one-week washout period, the same two groups underwent a single session of the other treatment, and the same measurements were taken. We measured walking speed, cadence, and the number of steps in a 10 m walking test (10MWT) and assessed Modified Ashworth Scale (MAS), Fugl-Meyer Assessment (FMA), and range of motion (ROM) before and after the intervention. RESULTS: Stepping + FES significantly improved walking speed, number of steps, and ankle inversion ROM, compared with stepping alone. Adverse events were not observed in any subject. CONCLUSIONS: Robotic stepping therapy combined with FES significantly improved 10 m walking speed (10MWS) compared with stepping only in patients with post-stroke and spastic hemiplegia. Further studies are needed to determine the long-term effects of the combination treatment.

Plectin promotes tumor formation by B16 mouse melanoma cells via regulation of Rous sarcoma oncogene activity.

BACKGROUND: Melanoma is a malignant tumor characterized by high proliferation and aggressive metastasis. To address the molecular mechanisms of the proto-oncogene, Rous sarcoma oncogene (Src), which is highly activated and promotes cell proliferation, migration, adhesion, and metastasis in melanoma. Plectin, a cytoskeletal protein, has recently been identified as a Src-binding protein that regulates Src activity in osteoclasts. Plectin is a candidate biomarker of certain tumors because of its high expression and the target of anti-tumor reagents such as ruthenium pyridinecarbothioamide. The molecular mechanisms by which plectin affects melanoma is still unclear. In this study, we examined the role of plectin in melanoma tumor formation. METHODS: We used CRISPR/Cas9 gene editing to knock-out plectin in B16 mouse melanoma cells. Protein levels of plectin and Src activity were examined by western blotting analysis. In vivo tumor formation was assessed by subcutaneous injection of B16 cells into nude mice and histological analysis performed after 2 weeks by Hematoxylin-Eosin (H&E) staining. Cell proliferation was evaluated by direct cell count, cell counting kit-8 assays, cyclin D1 mRNA expression and Ki-67 immunostaining. Cell aggregation and adhesion were examined by spheroid formation, dispase-based dissociation assay and cell adhesion assays. RESULTS: In in vivo tumor formation assays, depletion of plectin resulted in low-density tumors with large intercellular spaces. In vitro experiments revealed that plectin-deficient B16 cells exhibit reduced cell proliferation and reduced cell-to-cell adhesion. Since Src activity is reduced in plectin-deficient melanomas, we examined the relationship between plectin and Src signaling. Src overexpression in plectin knockout B16 cells rescued cell proliferation and improved cell-to-cell adhesion and cell to extracellular matrix adhesion. CONCLUSION: These results suggest that plectin plays critical roles in tumor formation by promoting cell proliferation and cell-to-cell adhesion through Src signaling activity in melanoma cells.

Analysis of external nasal morphology and the direction of maxillary repositioning following Le Fort I osteotomy.

This study investigated the relationship between the direction of maxillary repositioning after Le Fort I osteotomy and changes in external nasal morphology using lateral cephalograms and frontal facial photographs. The results indicated greater changes in external nasal morphology, such as a more forward position of the nasal tip and an increased alar base width, with anterosuperior repositioning than with posterosuperior repositioning. In conclusion, this study demonstrated that the external nasal morphology changes after Le Fort I osteotomy, and that the changes vary depending on the direction of maxilla repositioning.

Spinal Cord Injury in Middle-aged and Older Adults Who Had Undergone Active Rehabilitation Treatment at a Remote Hospital: A Case Series.

Background: Middle-aged and older individuals with spinal cord injury (SCI) often require long-term care even after receiving rehabilitation treatment, making it difficult for them to return home. We retrospectively investigated our active rehabilitation treatment for patients with SCI. Case: Included in this case series were ten patients with SCI who were admitted to our general hospital (located in the southern part of Wakayama Prefecture) and who underwent active rehabilitation treatment. The participants were investigated retrospectively by access to electronic medical records. The Barthel index scores for discharged patients were determined at an outpatient clinic, and the community phase of rehabilitation management was recorded. The average age of the 10 patients was 67.4 ± 13.4 years, and the average period from onset to transfer to our hospital was 102.6 ± 69.9 days. The Barthel index scores significantly improved from 39.0 ± 30.9 at admission to 65.0 ± 28.2 at discharge (P<0.05). Among the seven patients who were discharged to their homes, six had cervical SCI. Some patients with American Spinal Injury Association impairment scale grades A and B at admission could be discharged home, and their Barthel index scores were maintained after discharge. Discussion: : Even in a remote rural hospital, the activities of daily living of patients with SCI improved, and seven of the ten patients were discharged home. The activities of daily living of the discharged patients were maintained. To achieve these results, active rehabilitation treatment conducted by rehabilitation specialists is important.

Three-week exercise and protein intake immediately after exercise increases the 6-min walking distance with simultaneously improved plasma volume in patients with chronic cerebrovascular disease: a preliminary prospective study.

BACKGROUND: Blood volume (BV) is a critical factor for physical endurance in chronic stroke patients, while hypervolemia can worsen hypertension in these patients. This prospective study assessed whether rehabilitation combined with protein supplementation immediately after each exercise for 3 weeks would improve plasma volume (PV) and BV as well as physical endurance without worsening hypertension. METHODS: Ambulatory patients with chronic cerebrovascular disease who received a 3-week rehabilitation program with high protein jelly (intervention group [PG]; n = 8; 10-g protein) or protein-free jelly (control group [CG]; n = 8) consumed within 30 min after each exercise. PV and BV were assessed while measuring the 6-min walking distance (6MWD), peak oxygen consumption (VO2peak), strength of knee extension, and resting blood pressure before and after the intervention. Two-way ANOVA was used to determine whether there was an interaction of time × group. The difference between before and after intervention or between the groups by post-hoc test (Tukey's test) at the level of P < 0.05. RESULTS: The 6MWD increased only in the PG (P = 0.001; an interaction of Group and Time, P = 0.037). PV and BV increased only in the PG (P < 0.05). VO2peak and strength of knee extension in the paralysed limb increased in both groups (P < 0.05). The resting blood pressure did not worsen after the intervention. CONCLUSIONS: In chronic post-stroke patients, 3-week rehabilitation combined with protein intake immediately after exercise increased 6MWD simultaneously with increased PV and BV, but it did not increase resting blood pressure. The present regimen is acceptable and effective for ambulatory patients with chronic cerebrovascular disease. Name of the registry Examining effects of protein supplementation on functional improvement during rehabilitation intervention in chronic stroke patients Trial registration number UMIN000028009; date of registration: 30/06/2017. This study was registered prospectively.

Human telomerase reverse transcriptase protein expression is associated with survival in patients with oral squamous cell carcinoma.

Despite improvements in diagnosis and treatment, the 5-year survival of oral squamous cell carcinoma (OSCC), no matter the location, remains low, averaging 50%. Telomerase is expressed in 85% of malignancies and may play an important role in human carcinogenesis. Its catalytic component is human telomerase reverse transcriptase (hTERT), which has been thought, but not proven, to be involved in survival with OSCC. We investigated whether hTERT protein was a prognostic factor in OSCC by evaluating its association with clinicopathologic findings and OSCC survival. We found that in comparison to patients with high hTERT expression, patients with low hTERT expression survived significantly longer, including a longer 5-year overall survival. In addition, overall survival was significantly correlated to hTERT expression and the histologic grade and N status of the tumor. Disease-free survival was significantly related to hTERT expression, the histologic grade and N status of the tumor, and mode of invasion. These results suggest that hTERT protein is involved in cervical lymph node metastasis, that its levels may be increased during carcinogenesis, and that it may influence tumor invasion. We believe that this study is the first to demonstrate that OSCC with high hTERT expression carries a worse prognosis than cases with low hTERT expression.

Effective ancillary role and long-term course of daily or weekly teriparatide treatment on refractory medication-related osteonecrosis of the jaw: a clinical case series.

Medication-related osteonecrosis of the jaw (MRONJ) is a potentially severe adverse side effect of antiresorptive agents. However, withdrawal of such agents in patients with osteoporosis may increase the risk of fracture. The treatment of MRONJ is challenging, and standard treatment guidelines have yet to be established. In this study, the aim was to find out whether adjuvant daily or weekly teriparatide (TPTD) injections were beneficial for exposed bony MRONJ lesions compared with standard conservative management. We also studied the complications and the patients' response to TPTD therapy. We enrolled 27 patients (January 2012 - December 2016) with chronic and refractory MRONJ. There were four men and 23 women (85% female). Nine patients who did not select TPTD therapy for several reasons formed the non-TPTD group; the remaining 18 were randomly assigned to the daily (n=9) or weekly (n=9) groups. All patients in both groups continued standard conservative therapy in addition to their daily or weekly subcutaneous injection of TPTD (20 µg or 56.5 µg, respectively). We evaluated the complications of TPTD and its benefits. Three patients in the daily group did not complete the study, resulting in six patients in the daily group, nine in the weekly group, and nine in the non-TPTD group in the final analysis. The exposed bone was completely covered with normal mucosa in all patients in the TPTD groups, and the healing period was shorter than that in the non-TPTD group. No patient had complications of atypical fractures of the femoral head. Daily and weekly TPTD treatment resulted in a shortened treatment period compared with standard conservative therapy, with no increase in the rate of complications or worsening of osteoporosis.

Depth of invasion determined by magnetic resonance imaging in tongue cancer can be a predictor of cervical lymph node metastasis.

OBJECTIVES: We evaluated the relationships between depth of invasion (DOI) of tongue cancer, as measured with preoperative T1- and T2-weighted magnetic resonance imaging (MRI) and postoperative histopathologic (Path) specimens, with cervical lymph node metastasis (CLNM) and tumor stage. We also calculated the correlation of MRI and Path DOI measurements. STUDY DESIGN: This retrospective study included 101 patients who had squamous cell carcinoma of the tongue and were treated surgically. Two observers measured DOI on all 3 modalities. RESULTS: DOI thresholds for predicting CLNM with high diagnostic efficacy were 6.99 mm and 8.32 mm for MRI and 5 mm for Path. DOI values from all modalities were significantly different for tumors with and without CLNM (P < .01) and for the 4 TNM stages (P ≤ .05), with increasing values corresponding to advancement in tumor stage. Addition of DOI changed the T level of many tumors based on the new TNM (tumor-node-metastasis) classification. The correlation coefficient between DOI calculated on each MRI sequence and Path was 0.90. CONCLUSIONS: MRI-derived DOI accurately reflected the subsequent metastatic status and degree of progression of tumor stages, with a strong positive correlation to Path values, and may be considered a predictor of tumor stage and CLNM.

A simple sandwich technique using buttons combined with a tie-over technique for an intraoral split-thickness skin graft.

A simple sandwich technique using buttons to compress grafted tissue combined with a tie-over technique for intraoral split-thickness skin grafts (STSGs) is introduced. This technique yielded an excellent engraftment rate (90.5%) and no instances of total graft failure were recorded. This simple sandwich technique for STSGs is readily applicable and inexpensive, and the present results show that it would be potentially useful for repair of defects in the oral cavity.

Attenuation of core temperature elevation and interleukin-6 excretion during head-out hot water immersion in elderly people.

[Purpose] Previous studies have demonstrated a link between core body temperature and interleukin-6 production. Recent studies have reported that 20 minutes of head-out immersion in hot water (42°C) increased serum interleukin-6 levels in young males. This study aimed to compare the efficacy of head-out immersion in hot water (42°C) on serum interleukin-6 levels in seven elderly (66-75 years old) and eight young males (21-32 years old). [Participants and Methods] Venous blood samples were drawn at rest, immediately after head-out immersion in hot water (42°C), after 1 hour, and after 2 hours. Levels of serum interleukin-6, tumor necrosis factor-α, and high-sensitivity C-reactive protein; blood cell counts; and core temperature were measured. [Results] It was found that 20 minutes of head-out immersion in hot water (42°C) increased the core temperature in both the elderly and young participants; however, the rise in core temperature was more attenuated in elderly participants. Serum interleukin-6 levels were significantly higher in young participants 1 hour after the head-out immersion in hot water (42°C); however, serum interleukin-6 levels did not change in elderly participants. Serum tumor necrosis factor-α and high-sensitivity C-reactive protein levels remained constant throughout the study the elderly and young participants. [Conclusion] The current study demonstrated that head-out immersion in hot water (42°C) more attenuated core temperature and interleukin-6 levels in elderly participants than in young participants. We assert that these differences are likely to be related to age-related changes in core temperature regulation and muscle fibers.

p130Cas induces bone invasion by oral squamous cell carcinoma by regulating tumor epithelial-mesenchymal transition and cell proliferation.

Bone invasion is a critical factor in determining the prognosis of oral squamous cell carcinoma (OSCC) patients. Transforming growth factor ß (TGF-ß) is abundantly expressed in the bone matrix and is involved in the acquisition of aggressiveness by tumors. TGF-ß is also important to cytoskeletal changes during tumor progression. In this study, we examined the relationship between TGF-ß signaling and cytoskeletal changes during bone invasion by OSCC. Immunohistochemical staining of OSCC samples from five patients showed the expression of p130Cas (Crk-associated substrate) in the cytoplasm and phosphorylated Smad3 expression in the nucleus in OSCC cells. TGF-ß1 induced the phosphorylation of Smad3 and p130Cas, as well as epithelial-mesenchymal transition (EMT) accompanied by the downregulation of the expression of E-cadherin, a marker of epithelial cells, and the upregulation of the expression of N-cadherin, or Snail, a marker of mesenchymal cells, in human HSC-2 cells and mouse squamous cell carcinome VII (SCCVII) cells. SB431542, a specific inhibitor of Smad2/3 signaling, abrogated the TGF-ß1-induced phosphorylation of p130Cas and morphological changes. Silencing p130Cas using an short hairpin RNA (shRNA) or small interfering RNA in SCCVII cells suppressed TGF-ß1-induced cell migration, invasion, EMT and matrix metalloproteinase-9 (MMP-9) production. Compared with control SCCVII cells, SCCVII cells with silenced p130Cas strongly suppressed zygomatic and mandibular destruction in vivo by reducing the number of osteoclasts, cell proliferation and MMP-9 production. Taken together, these results showed that the expression of TGF-ß/p130Cas might be a new target for the treatment of OSCC bone invasion.

Plectin stabilizes microtubules during osteoclastic bone resorption by acting as a scaffold for Src and Pyk2.

Osteoclasts are multinuclear cells which maintain bone homeostasis by resorbing bone. During bone resorption, osteoclasts attach to the bone matrix via a sealing zone formed by an actin ring. Rous sarcoma oncogene (Src) is essential for actin ring formation and bone resorption. Recently, we demonstrated that plectin, a cytolinker protein, is a Src-binding protein in osteoclasts. However, the function of plectin in osteoclasts remains unknown. In this study, we demonstrated that shRNA knockdown of plectin in RAW 264.7 cells resulted in tartrate resistant acid phosphatase positive multinuclear cells (TRAP (+) MNCs) with impaired actin ring formation and bone resorption activity. Moreover, we found that in plectin-silenced TRAP (+) MNCs, Src and protein tyrosine kinase 2 beta (Pyk2), two critical kinases in osteoclastic bone resorption, were inactivated and microtubule polarity was disturbed. These results suggest that plectin plays a critical role in osteoclast biology by acting as a scaffold to facilitate Src and Pyk2 activation during microtubule organization.

The Biological Effects of Interleukin-17A on Adhesion Molecules Expression and Foam Cell Formation in Atherosclerotic Lesions.

Interleukin-17A (IL-17A), a major effector cytokine secreted by T helper 17 (Th17) cells, is elevated in atherosclerosis lesions. The purpose of this study was to assess the role of IL-17A in the pathogenesis of atherosclerosis. To measure the expression of adhesion molecules, human umbilical vein endothelial cells (HUVECs) and U937 cells were stimulated with IL-17A. Western blot and real-time polymerase chain reaction analyses revealed that intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) expression in HUVECs, and very late antigen-4 (VLA-4), lymphocyte function-associated antigen-1 (LFA-1) and macrophage-1 antigen (MAC-1) expression in U937 cells was upregulated by IL-17A. Furthermore, IL-17A stimulation resulted in mRNA and protein expression of scavenger receptor (LOX-1) in phorbol 12-myristate 13-acetate (PMA)-activated U937 cells. Oil Red O also demonstrated that IL-17A enhanced foam cell formation by PMA-activated U937 cells induced by oxidized low-density lipoprotein (ox-LDL), and this enhancement of ox-LDL-induced foam cell formation in IL-17A-treated U937 cells was downregulated by transfection of LOX-1 siRNA. These results indicated that IL-17A induced the expression of adhesion molecules, promoted the adherence of monocytes to vascular endothelial cells. IL-17A also stimulated ox-LDL-induced foam cell formation via upregulation of LOX-1 in activated macrophages. IL-17A may be responsible for the pathogenesis of atherosclerosis by inducing the adhesion of leukocytes to vascular endothelium and foam cell formation.

Accumulation of hyaluronic acid in stromal cells modulates osteoclast formation by regulation of receptor activator of nuclear factor kappa-B ligand expression.

Hyaluronic acid (HA) has a pivotal role in bone and cartilage metabolism. In this study, we investigated the effect and underlying mechanisms of HA accumulation on the expression of receptor activator of nuclear factor kappa-B ligand (RANKL) induced by 1α,25(OH)2D3 and dexamethasone in stromal cells, which support osteoclastogenesis. Degradation of HA by hyaluronidase (HA'ase) treatment enhanced the expression of RANKL in ST2 cells stimulated with 1α,25(OH)2D3 and dexamethasone. Down-regulation of hyaluronan synthase 2 (HAS2) expression by siRNA also stimulated RANKL expression induced by 1α,25(OH)2D3 and dexamethasone. Results from a cell co-culture system with bone marrow cell showed that 1α,25(OH)2D3 and dexamethasone-induced RANKL expression in HA'ase treated- and HAS2 siRNA transfected-ST2 cells was down-regulated by treatment of cells with high molecular weight HA. In contrast, transforming growth factor-ß1 (TGF-ß1), which stimulates HAS2 expression and HA synthesis, down-regulated RANKL expression induced by 1α,25(OH)2D3 and dexamethasone. Interestingly, knockdown of has2 gene enhanced the expression of vitamin D receptor (VDR) and phosphorylation of signal transducers and activator of transcription 3 (STAT3) in ST2 cells stimulated by 1α,25(OH)2D3 and dexamethasone. These results indicate that accumulation of HA in bone marrow cells may affect RANKL-mediated osteoclast-supporting activity via regulation of VDR and STAT3 signaling pathways.